lipofectamine 2000  (Thermo Fisher)


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    Name:
    Lipofectamine 2000 Transfection Reagent
    Description:
    Lipofectamine 2000 Transfection Reagent is a versatile transfection reagent that has been shown to effectively transfect the widest variety of adherent and suspension cell lines Researchers use Lipofectamine 2000 Reagent for siRNA and shRNA based gene knockdown experiments as well as for gene expression studies With Lipofectamine 2000 Transfection Reagent you ll get • Exceptional transfection efficiency in the broadest range of cell lines and the highest levels of recombinant protein expression see table • Superior performance for co transfection of siRNA and plasmid DNA• Proven efficacy in the presence of serum eliminates the need to change media following transfection• Reliable performance for high throughput applications• The best choice for establishing stable cell linesA high performance transfection reagent for gene expression and gene silencingLipofectamine 2000 Transfection Reagent works effectively with all common cell lines as well as with many challenging ones and can be used in media with or without serum For gene silencing Lipofectamine 2000 Transfection Reagent s high efficiency transfections provide the high levels of gene knockdown needed to produce convincing results Lipofectamine 2000 Transfection Reagent is the number one choice for co transfection given its effectiveness for transfecting both siRNA and plasmid DNA Lipofectamine 2000 Transfection Reagent is easy to use simply mix with nucleic acid and add to cell culture Ideal for high throughput workSimplicity and speed combined with high transfection efficiency make Lipofectamine 2000 Transfection Reagent ideal for transient protein expression or high throughput RNAi transfections Transfection conditions can be easily established for automated or robotic systems used in such applications
    Catalog Number:
    11668019
    Price:
    None
    Category:
    Cell Culture Transfection Reagents
    Applications:
    Cell Culture|Plasmid Transfection|RNAi Transfection|Stem Cell & Primary Cell Transfections|Synthetic siRNA Transfection|shRNA & miR RNAi Plasmid Transfection|Transfection
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    Structured Review

    Thermo Fisher lipofectamine 2000
    Evaluation of G4Arg-based transfection in MEFs. Notes:  ( A ) Cytotoxicity of G4Arg nanoparticles at 24 hours and 48 hours as assessed by MTT assay. ( B ) The fluorescence expressed from MEFs after pOSKMG delivery based on G4Arg, Lipofectamine ®  2000, and FuGENE ®  vectors. Scale bar =100 μm. ( C ) G4Arg-mediated transfection efficiency at various N/P ratios. ( D ) G4Arg-mediated transfection efficiency using various plasmid dosages. ( E ) G4Arg-mediated transfection efficiency using various transfection times. ( F ) Comparison of transfection efficiency based on G4Arg, Lipofectamine 2000, and FuGENE vectors (*versus G4Arg,  P
    Lipofectamine 2000 Transfection Reagent is a versatile transfection reagent that has been shown to effectively transfect the widest variety of adherent and suspension cell lines Researchers use Lipofectamine 2000 Reagent for siRNA and shRNA based gene knockdown experiments as well as for gene expression studies With Lipofectamine 2000 Transfection Reagent you ll get • Exceptional transfection efficiency in the broadest range of cell lines and the highest levels of recombinant protein expression see table • Superior performance for co transfection of siRNA and plasmid DNA• Proven efficacy in the presence of serum eliminates the need to change media following transfection• Reliable performance for high throughput applications• The best choice for establishing stable cell linesA high performance transfection reagent for gene expression and gene silencingLipofectamine 2000 Transfection Reagent works effectively with all common cell lines as well as with many challenging ones and can be used in media with or without serum For gene silencing Lipofectamine 2000 Transfection Reagent s high efficiency transfections provide the high levels of gene knockdown needed to produce convincing results Lipofectamine 2000 Transfection Reagent is the number one choice for co transfection given its effectiveness for transfecting both siRNA and plasmid DNA Lipofectamine 2000 Transfection Reagent is easy to use simply mix with nucleic acid and add to cell culture Ideal for high throughput workSimplicity and speed combined with high transfection efficiency make Lipofectamine 2000 Transfection Reagent ideal for transient protein expression or high throughput RNAi transfections Transfection conditions can be easily established for automated or robotic systems used in such applications
    https://www.bioz.com/result/lipofectamine 2000/product/Thermo Fisher
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    lipofectamine 2000 - by Bioz Stars, 2021-07
    99/100 stars

    Images

    1) Product Images from "Reprogramming fibroblasts to pluripotency using arginine-terminated polyamidoamine nanoparticles based non-viral gene delivery system"

    Article Title: Reprogramming fibroblasts to pluripotency using arginine-terminated polyamidoamine nanoparticles based non-viral gene delivery system

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S73961

    Evaluation of G4Arg-based transfection in MEFs. Notes:  ( A ) Cytotoxicity of G4Arg nanoparticles at 24 hours and 48 hours as assessed by MTT assay. ( B ) The fluorescence expressed from MEFs after pOSKMG delivery based on G4Arg, Lipofectamine ®  2000, and FuGENE ®  vectors. Scale bar =100 μm. ( C ) G4Arg-mediated transfection efficiency at various N/P ratios. ( D ) G4Arg-mediated transfection efficiency using various plasmid dosages. ( E ) G4Arg-mediated transfection efficiency using various transfection times. ( F ) Comparison of transfection efficiency based on G4Arg, Lipofectamine 2000, and FuGENE vectors (*versus G4Arg,  P
    Figure Legend Snippet: Evaluation of G4Arg-based transfection in MEFs. Notes: ( A ) Cytotoxicity of G4Arg nanoparticles at 24 hours and 48 hours as assessed by MTT assay. ( B ) The fluorescence expressed from MEFs after pOSKMG delivery based on G4Arg, Lipofectamine ® 2000, and FuGENE ® vectors. Scale bar =100 μm. ( C ) G4Arg-mediated transfection efficiency at various N/P ratios. ( D ) G4Arg-mediated transfection efficiency using various plasmid dosages. ( E ) G4Arg-mediated transfection efficiency using various transfection times. ( F ) Comparison of transfection efficiency based on G4Arg, Lipofectamine 2000, and FuGENE vectors (*versus G4Arg, P

    Techniques Used: Transfection, MTT Assay, Fluorescence, Plasmid Preparation

    2) Product Images from "A combinatorial approach to determine functional group effects on lipidoid-mediated siRNA delivery"

    Article Title: A combinatorial approach to determine functional group effects on lipidoid-mediated siRNA delivery

    Journal: Bioconjugate chemistry

    doi: 10.1021/bc100041r

    Viability of HeLa cells post-transfection with selected lipidoids. HeLa cells were transfected with 50 ng siRNA complexed at lipidoid/ siRNA ratios of 2.5:1, 5:1, 10:1 and 15:1 (wt/wt). Lipofectamine 2000 was used as instructed by the vendor. After incubation
    Figure Legend Snippet: Viability of HeLa cells post-transfection with selected lipidoids. HeLa cells were transfected with 50 ng siRNA complexed at lipidoid/ siRNA ratios of 2.5:1, 5:1, 10:1 and 15:1 (wt/wt). Lipofectamine 2000 was used as instructed by the vendor. After incubation

    Techniques Used: Transfection, Incubation

    Library screening for siRNA transfection efficiency. HeLa cells stably expressing firefly and Renilla luciferase were transfected with 50 ng of firefly specific siRNA complexed at lipidoid/ siRNA ratios of 2.5:1, 5:1, 10:1 and 15:1 (wt/wt). Lipofectamine
    Figure Legend Snippet: Library screening for siRNA transfection efficiency. HeLa cells stably expressing firefly and Renilla luciferase were transfected with 50 ng of firefly specific siRNA complexed at lipidoid/ siRNA ratios of 2.5:1, 5:1, 10:1 and 15:1 (wt/wt). Lipofectamine

    Techniques Used: Library Screening, Transfection, Stable Transfection, Expressing, Luciferase

    3) Product Images from "KDM5 histone demethylases repress immune response via suppression of STING"

    Article Title: KDM5 histone demethylases repress immune response via suppression of STING

    Journal: PLoS Biology

    doi: 10.1371/journal.pbio.2006134

    KDM5 inhibition induces a robust interferon response in cancer cells with elevated levels of cytosolic DNA. (A) Immunostaining of dsDNA and the mitochondrial marker Hsp60 in MCF7 cells. Surface plots of Z-stack images generated with Huygens (left panel). qPCR detecting mtDNA copy number in cells with the indicated treatment (right panel). 10 μM of ddC was used. Scale bar, 10 μm. (B, C) RT-qPCR (panel B) and western blot (panel C) analyses of MCF7 cells with the indicated treatment. MCF7 cells were treated with 10 μM ddC and 1 μM KDM5-C70 for 6 days. Cells were refed every 2 days. (D) dsDNA and DAPI staining of MCF7 cells or SKBR3 cells digested with 50 μg/ml DNase I. Surface plots of Z-stack images generated with Huygens (left panel). Quantification of dsDNA intensity per cell using image J was shown in the right panel. Scale bar, 10 μm. (E) RT-qPCR analyses of SKBR3 cells with the indicated treatment. SKBR3 cells were transfected with 1 μg dsDNA of about 90 bp (dsDNA90) using Lipofectamine 2000 5 hours before treatment with DMSO or 1 μM KDM5-C70 for 3 days. (F) RT-qPCR analysis of SKBR3 cells with knockout of the indicated genes after treatment with 1 μg dsDNA and 1 μM KDM5-C70 for 3 days. Representative data from triplicate experiments are shown. Error bar denotes SEM. # p
    Figure Legend Snippet: KDM5 inhibition induces a robust interferon response in cancer cells with elevated levels of cytosolic DNA. (A) Immunostaining of dsDNA and the mitochondrial marker Hsp60 in MCF7 cells. Surface plots of Z-stack images generated with Huygens (left panel). qPCR detecting mtDNA copy number in cells with the indicated treatment (right panel). 10 μM of ddC was used. Scale bar, 10 μm. (B, C) RT-qPCR (panel B) and western blot (panel C) analyses of MCF7 cells with the indicated treatment. MCF7 cells were treated with 10 μM ddC and 1 μM KDM5-C70 for 6 days. Cells were refed every 2 days. (D) dsDNA and DAPI staining of MCF7 cells or SKBR3 cells digested with 50 μg/ml DNase I. Surface plots of Z-stack images generated with Huygens (left panel). Quantification of dsDNA intensity per cell using image J was shown in the right panel. Scale bar, 10 μm. (E) RT-qPCR analyses of SKBR3 cells with the indicated treatment. SKBR3 cells were transfected with 1 μg dsDNA of about 90 bp (dsDNA90) using Lipofectamine 2000 5 hours before treatment with DMSO or 1 μM KDM5-C70 for 3 days. (F) RT-qPCR analysis of SKBR3 cells with knockout of the indicated genes after treatment with 1 μg dsDNA and 1 μM KDM5-C70 for 3 days. Representative data from triplicate experiments are shown. Error bar denotes SEM. # p

    Techniques Used: Inhibition, Immunostaining, Marker, Generated, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Staining, Transfection, Knock-Out

    4) Product Images from "Enhanced mRNA delivery into lymphocytes enabled by lipid-varied libraries of charge-altering releasable transporters"

    Article Title: Enhanced mRNA delivery into lymphocytes enabled by lipid-varied libraries of charge-altering releasable transporters

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.1805358115

    ( A ) Structure of previously reported CART D 13 :A 11 and neutral products of the charge-altering mechanism. ( B ) Transfection efficiencies of EGFP mRNA alone, lipofectamine-formulated EGFP mRNA, and D 13 :A 11 -complexed EGFP mRNA into HeLa cells versus Jurkat cells (T lymphocytes). Error bars represent the SD over three experiments in triplicate.
    Figure Legend Snippet: ( A ) Structure of previously reported CART D 13 :A 11 and neutral products of the charge-altering mechanism. ( B ) Transfection efficiencies of EGFP mRNA alone, lipofectamine-formulated EGFP mRNA, and D 13 :A 11 -complexed EGFP mRNA into HeLa cells versus Jurkat cells (T lymphocytes). Error bars represent the SD over three experiments in triplicate.

    Techniques Used: Transfection

    5) Product Images from "Innovative approaches to genome editing in avian species"

    Article Title: Innovative approaches to genome editing in avian species

    Journal: Journal of Animal Science and Biotechnology

    doi: 10.1186/s40104-018-0231-7

    Chicken, turkey, and quail sperm incubated with Lipofectamine® 2000 and BLOCK-iT™ fluorescently labelled RNA. The top panel shows unprocessed sperm, where poor transfection of the labelled RNA to the sperm is seen. The bottom panel shows STAGE processed sperm, where these is clearly increased transfection of the labelled RNA to the sperm. Quail pictures taken by Olivier Serralbo of the Monash Transgenic Quail Facility
    Figure Legend Snippet: Chicken, turkey, and quail sperm incubated with Lipofectamine® 2000 and BLOCK-iT™ fluorescently labelled RNA. The top panel shows unprocessed sperm, where poor transfection of the labelled RNA to the sperm is seen. The bottom panel shows STAGE processed sperm, where these is clearly increased transfection of the labelled RNA to the sperm. Quail pictures taken by Olivier Serralbo of the Monash Transgenic Quail Facility

    Techniques Used: Incubation, Blocking Assay, Transfection, Transgenic Assay

    6) Product Images from "VBP1 facilitates proteasome and autophagy-mediated degradation of MutS homologue hMSH4"

    Article Title: VBP1 facilitates proteasome and autophagy-mediated degradation of MutS homologue hMSH4

    Journal: The FASEB Journal

    doi: 10.1096/fj.13-235127

    hMSH4 interacts with the VBP1-VHL-p97 complex. A ) Interaction between hMSH4 and VBP1 was analyzed in HEK293T cells by reciprocal coimmunoprecipitation (co-IP) as indicated. B ) VHL and p97 coexist in the hMSH4 immunocomplex. HEK293T cells transfected with empty vector or Myc-hMSH4 were subjected to immunoprecipitation using an anti-Myc antibody. C ) VBP1 modulates the interaction between hMSH4 and p97. HEK293T cells were first transfected with control shRNA or VBP1 shRNA sh-2 to knock down VBP1 and were then transfected with Myc-hMSH4. At 24 h post-transfection, cells were treated with MG-132 and subjected to immunoprecipitation using an anti-Myc antibody, and immunoblotting was carried out as indicated. D ) HEK293T cells were transfected with p97 siRNA using Lipofectamine 2000 before transfection with Myc-hMSH4. The protein level of hMSH4 was examined by immunoblotting.
    Figure Legend Snippet: hMSH4 interacts with the VBP1-VHL-p97 complex. A ) Interaction between hMSH4 and VBP1 was analyzed in HEK293T cells by reciprocal coimmunoprecipitation (co-IP) as indicated. B ) VHL and p97 coexist in the hMSH4 immunocomplex. HEK293T cells transfected with empty vector or Myc-hMSH4 were subjected to immunoprecipitation using an anti-Myc antibody. C ) VBP1 modulates the interaction between hMSH4 and p97. HEK293T cells were first transfected with control shRNA or VBP1 shRNA sh-2 to knock down VBP1 and were then transfected with Myc-hMSH4. At 24 h post-transfection, cells were treated with MG-132 and subjected to immunoprecipitation using an anti-Myc antibody, and immunoblotting was carried out as indicated. D ) HEK293T cells were transfected with p97 siRNA using Lipofectamine 2000 before transfection with Myc-hMSH4. The protein level of hMSH4 was examined by immunoblotting.

    Techniques Used: Co-Immunoprecipitation Assay, Transfection, Plasmid Preparation, Immunoprecipitation, shRNA

    7) Product Images from "RING-finger protein 166 plays a novel pro-apoptotic role in neurotoxin-induced neurodegeneration via ubiquitination of XIAP"

    Article Title: RING-finger protein 166 plays a novel pro-apoptotic role in neurotoxin-induced neurodegeneration via ubiquitination of XIAP

    Journal: Cell Death & Disease

    doi: 10.1038/s41419-020-03145-x

    Pro-apoptotic function of RNF166 in naturally occurring neuronal death. a – b Primary hippocampal neurons at DIV 1 were transiently transfected with GFP- or GFP-tagged RNF166 using Lipofectamine 2000 and further cultivated for the indicated time periods. a At the indicated time periods, cells were counterstained with 1 mg/mL Hoechst 33258 and then examined under confocal microscopy. Representative fluorescence images are shown. Arrowheads indicate typical dying cells with no apparent GFP signal or fragmented GFP staining. Scale bar, 50 µm. GFP-positive cells were counted by flow cytometry analysis. Values are expressed as the number of GFP-positive cells at the indicated time period after transfection. Each value represents the mean + SEM of at least three separate experiments. **** p
    Figure Legend Snippet: Pro-apoptotic function of RNF166 in naturally occurring neuronal death. a – b Primary hippocampal neurons at DIV 1 were transiently transfected with GFP- or GFP-tagged RNF166 using Lipofectamine 2000 and further cultivated for the indicated time periods. a At the indicated time periods, cells were counterstained with 1 mg/mL Hoechst 33258 and then examined under confocal microscopy. Representative fluorescence images are shown. Arrowheads indicate typical dying cells with no apparent GFP signal or fragmented GFP staining. Scale bar, 50 µm. GFP-positive cells were counted by flow cytometry analysis. Values are expressed as the number of GFP-positive cells at the indicated time period after transfection. Each value represents the mean + SEM of at least three separate experiments. **** p

    Techniques Used: Transfection, Confocal Microscopy, Fluorescence, Staining, Flow Cytometry

    8) Product Images from "Effect of celastrol on toll-like receptor 4-mediated inflammatory response in free fatty acid-induced HepG2 cells"

    Article Title: Effect of celastrol on toll-like receptor 4-mediated inflammatory response in free fatty acid-induced HepG2 cells

    Journal: International Journal of Molecular Medicine

    doi: 10.3892/ijmm.2018.3775

    Effects of transfection with TLR4 siRNA in the HepG2 cells. In order to select the most effective TLR4 siRNA sequences, three FAM-labeled TLR4 siRNA constructs were transiently transfected into HepG2 cells using Lipofectamine ®  2000. After 24 h of transfection, the transfection efficiency was tested by a FAM-siRNA under a fluorescence microscope (magnification, x200). (A) Visualization of the cells under common light and FAM-fluorescence light. The fluorescent particles within the cells indicated that siRNA was transfected successfully into the HepG2 cells. (B) Relative mRNA and (C) relative protein expression levels of TLR4 were analyzed by reverse transcription-quantitative polymerase chain and western blotting, respectively. (D) FFA + negative control siRNA and (E) TLR4 siRNA-treated HepG2 cells stained by Oil red O (magnification, x400). Red arrows indicate lipid droplets. (F) Levels of intracellular triglycerides were analyzed by an enzymatic assay. Data are expressed as the mean ± standard error of the mean (n=6).  * P
    Figure Legend Snippet: Effects of transfection with TLR4 siRNA in the HepG2 cells. In order to select the most effective TLR4 siRNA sequences, three FAM-labeled TLR4 siRNA constructs were transiently transfected into HepG2 cells using Lipofectamine ® 2000. After 24 h of transfection, the transfection efficiency was tested by a FAM-siRNA under a fluorescence microscope (magnification, x200). (A) Visualization of the cells under common light and FAM-fluorescence light. The fluorescent particles within the cells indicated that siRNA was transfected successfully into the HepG2 cells. (B) Relative mRNA and (C) relative protein expression levels of TLR4 were analyzed by reverse transcription-quantitative polymerase chain and western blotting, respectively. (D) FFA + negative control siRNA and (E) TLR4 siRNA-treated HepG2 cells stained by Oil red O (magnification, x400). Red arrows indicate lipid droplets. (F) Levels of intracellular triglycerides were analyzed by an enzymatic assay. Data are expressed as the mean ± standard error of the mean (n=6). * P

    Techniques Used: Transfection, Labeling, Construct, Fluorescence, Microscopy, Expressing, Western Blot, Negative Control, Staining, Enzymatic Assay

    9) Product Images from "Targeted therapy against Bcl-2-related proteins in breast cancer cells"

    Article Title: Targeted therapy against Bcl-2-related proteins in breast cancer cells

    Journal: Breast Cancer Research

    doi: 10.1186/bcr1323

    Effect of treatment of ZR-75-1 cells with antisense Bcl-2 and doxorubicin on the apoptosis-related proteins. Cells were pretreated with 10 μg/ml Lipofectamine alone, or 1.0 μM antisense Bcl-2 oligodeoxynucleotides, for 24 hours. Cells were then cultured in standard medium, after which they were treated with 0.5 μM doxorubicin (DOX) for 6, 12, or 24 hours. Whole cell lysate was then extracted and subjected to Western blotting. The data presented are from more than two independent experiments.
    Figure Legend Snippet: Effect of treatment of ZR-75-1 cells with antisense Bcl-2 and doxorubicin on the apoptosis-related proteins. Cells were pretreated with 10 μg/ml Lipofectamine alone, or 1.0 μM antisense Bcl-2 oligodeoxynucleotides, for 24 hours. Cells were then cultured in standard medium, after which they were treated with 0.5 μM doxorubicin (DOX) for 6, 12, or 24 hours. Whole cell lysate was then extracted and subjected to Western blotting. The data presented are from more than two independent experiments.

    Techniques Used: Cell Culture, Western Blot

    Sequence-specific downregulation and cytotoxic effects of antisense Bcl-2 oligodeoxynucleotides on BT-474 and ZR-75-1 cells. (a) Specific inhibition of Bcl-2 protein expression by treatment with antisense (AS) Bcl-2 oligodeoxynucleotides (ODNs). Cells were treated with 10 μg/ml Lipofectamine alone (control) or 1.0 μM AS, mismatch control (MM), or random control (RC) ODNs for 24 hours. Cells were then cultured in standard medium, total protein was extracted, and Bcl-2 and β-actin protein levels were analyzed by Western blotting. (b) Quantification of Bcl-2 expression by densitometric analysis. The expression of Bcl-2 in cells treated with control, AS Bcl-2 , RC Bcl-2 , and MM Bcl-2 ODNs was normalized with β-actin, and the relative values are presented. Error bars indicate SD. The data presented are from three independent experiments. (c) Effects of AS Bcl-2 ODNs on the proliferation of BT-474 and ZR-75-1 breast cancer cells in vitro . Cells were treated with various concentrations of AS Bcl-2 ODNs in 24-well dishes. Four days after treatment, cells were stained with trypan blue and counted. Error bars indicate SD. The data presented are from three independent experiments.
    Figure Legend Snippet: Sequence-specific downregulation and cytotoxic effects of antisense Bcl-2 oligodeoxynucleotides on BT-474 and ZR-75-1 cells. (a) Specific inhibition of Bcl-2 protein expression by treatment with antisense (AS) Bcl-2 oligodeoxynucleotides (ODNs). Cells were treated with 10 μg/ml Lipofectamine alone (control) or 1.0 μM AS, mismatch control (MM), or random control (RC) ODNs for 24 hours. Cells were then cultured in standard medium, total protein was extracted, and Bcl-2 and β-actin protein levels were analyzed by Western blotting. (b) Quantification of Bcl-2 expression by densitometric analysis. The expression of Bcl-2 in cells treated with control, AS Bcl-2 , RC Bcl-2 , and MM Bcl-2 ODNs was normalized with β-actin, and the relative values are presented. Error bars indicate SD. The data presented are from three independent experiments. (c) Effects of AS Bcl-2 ODNs on the proliferation of BT-474 and ZR-75-1 breast cancer cells in vitro . Cells were treated with various concentrations of AS Bcl-2 ODNs in 24-well dishes. Four days after treatment, cells were stained with trypan blue and counted. Error bars indicate SD. The data presented are from three independent experiments.

    Techniques Used: Sequencing, Inhibition, Expressing, Cell Culture, Western Blot, In Vitro, Staining

    Sequence-specific downregulation and cytotoxic effects of antisense Bcl-xL oligodeoxynucleotides on MDA-MB-231 and BT-474 cells. (a) Specific inhibition of Bcl-xL protein expression by treatment with antisense (AS) Bcl-xL oligodeoxynucleotides (ODNs). Cells were treated with 10 μg/ml Lipofectamine alone (control) or 1.0 μM AS, mismatch control (MM), or random control (RC) ODNs for 24 hours. Cells were then cultured in standard medium, total protein was extracted, and Bcl-xL and β-actin protein levels were analyzed by Western blotting. (b) Quantification of Bcl-xL protein expression by densitometric analysis. The Bcl-xL protein expression was normalized with β-actin, and the relative values are presented. Error bars indicate SD. The data presented are from three independent experiments. (c) Effects of AS Bcl-xL ODNs on the proliferation of MDA-MB-231 and BT-474 breast cancer cells in vitro . Cells were treated with various concentrations of AS Bcl-xL ODNs in 24-well dishes. Four days after treatment, cells were stained with trypan blue and counted. Error bars indicate SD. The data presented are from three independent experiments.
    Figure Legend Snippet: Sequence-specific downregulation and cytotoxic effects of antisense Bcl-xL oligodeoxynucleotides on MDA-MB-231 and BT-474 cells. (a) Specific inhibition of Bcl-xL protein expression by treatment with antisense (AS) Bcl-xL oligodeoxynucleotides (ODNs). Cells were treated with 10 μg/ml Lipofectamine alone (control) or 1.0 μM AS, mismatch control (MM), or random control (RC) ODNs for 24 hours. Cells were then cultured in standard medium, total protein was extracted, and Bcl-xL and β-actin protein levels were analyzed by Western blotting. (b) Quantification of Bcl-xL protein expression by densitometric analysis. The Bcl-xL protein expression was normalized with β-actin, and the relative values are presented. Error bars indicate SD. The data presented are from three independent experiments. (c) Effects of AS Bcl-xL ODNs on the proliferation of MDA-MB-231 and BT-474 breast cancer cells in vitro . Cells were treated with various concentrations of AS Bcl-xL ODNs in 24-well dishes. Four days after treatment, cells were stained with trypan blue and counted. Error bars indicate SD. The data presented are from three independent experiments.

    Techniques Used: Sequencing, Multiple Displacement Amplification, Inhibition, Expressing, Cell Culture, Western Blot, In Vitro, Staining

    10) Product Images from "Self-complementarity in adeno-associated virus enhances transduction and gene expression in mouse cochlear tissues"

    Article Title: Self-complementarity in adeno-associated virus enhances transduction and gene expression in mouse cochlear tissues

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0242599

    Self-complementarity increased transduction efficiency of AAV2-mediated GFP expression in the organ of Corti. (A) A representative image from a neonatal cochlear explant treated with ssAAV2-CMV-GFP (SS-Ctrl) with GFP shown in green and parvalbumin (PVALB) shown in magenta. (B) A representative image from a neonatal cochlear explant treated with scAAV2-CMV-GFP which shows a large number of cells in the organ of Corti as being GFP positive. (C–E) Show images from explants treated with ssAAV2-CMV-GFP virions that were either packaged with Lipofectamine 2000 reagent (C) or pretreated with EERI (D) or Tyr23 (E). Post-hoc comparisons of each treatment group to the control group showed a significant effect of self-complementarity (F) on total numbers of GFP positive cells in the organ of Corti, and similarly a significant effect of self-complementary AAV on the numbers of GFP positive hair cells specifically (G). Error bars represent ±1 SEM, *** denotes p
    Figure Legend Snippet: Self-complementarity increased transduction efficiency of AAV2-mediated GFP expression in the organ of Corti. (A) A representative image from a neonatal cochlear explant treated with ssAAV2-CMV-GFP (SS-Ctrl) with GFP shown in green and parvalbumin (PVALB) shown in magenta. (B) A representative image from a neonatal cochlear explant treated with scAAV2-CMV-GFP which shows a large number of cells in the organ of Corti as being GFP positive. (C–E) Show images from explants treated with ssAAV2-CMV-GFP virions that were either packaged with Lipofectamine 2000 reagent (C) or pretreated with EERI (D) or Tyr23 (E). Post-hoc comparisons of each treatment group to the control group showed a significant effect of self-complementarity (F) on total numbers of GFP positive cells in the organ of Corti, and similarly a significant effect of self-complementary AAV on the numbers of GFP positive hair cells specifically (G). Error bars represent ±1 SEM, *** denotes p

    Techniques Used: Transduction, Expressing

    Self-complementarity increased transduction efficiency of AAV1-mediated GFP expression in the organ of Corti. (A) A representative image from a neonatal cochlear explant treated with ssAAV1_CMV_GFP (SS-Ctrl) with GFP shown in green and parvalbumin (PVALB) shown in magenta. (B) A representative image from a neonatal cochlear explant treated with scAAV1-CMV-GFP. (C–E) Show images from explants treated with ssAAV1-CMV-GFP virions that were either packaged with Lipofectamine 2000 reagent (C) or pretreated with EERI (D) or Tyr23 (E). Post-hoc comparisons of each treatment group to the control showed a significant effect of self-complementarity (F) on total numbers of GFP positive cells in the organ of Corti, but no effect of any treatment condition on the numbers of GFP positive hair cells (G). Error bars represent 1 SEM, * denotes p
    Figure Legend Snippet: Self-complementarity increased transduction efficiency of AAV1-mediated GFP expression in the organ of Corti. (A) A representative image from a neonatal cochlear explant treated with ssAAV1_CMV_GFP (SS-Ctrl) with GFP shown in green and parvalbumin (PVALB) shown in magenta. (B) A representative image from a neonatal cochlear explant treated with scAAV1-CMV-GFP. (C–E) Show images from explants treated with ssAAV1-CMV-GFP virions that were either packaged with Lipofectamine 2000 reagent (C) or pretreated with EERI (D) or Tyr23 (E). Post-hoc comparisons of each treatment group to the control showed a significant effect of self-complementarity (F) on total numbers of GFP positive cells in the organ of Corti, but no effect of any treatment condition on the numbers of GFP positive hair cells (G). Error bars represent 1 SEM, * denotes p

    Techniques Used: Transduction, Expressing

    11) Product Images from "Triolein-based polycation lipid nanocarrier for efficient gene delivery: characteristics and mechanism"

    Article Title: Triolein-based polycation lipid nanocarrier for efficient gene delivery: characteristics and mechanism

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S24720

    Transfection efficiency of polycation lipid nanocarrier/DNA complexes (PDC) (N/P = 10) in human lung adenocarcinoma (SPC-A1) cells compared with those of Lipofectamine™ 2000/DNA complexes (LDC). ( A ) Transfection efficiency of plasmid pEGFP-N2 determined by flow cytometer; ( B ) relative luciferase activity in SPC-A1 cells treated with PDC in comparison with that of LDC; ( C ) the cell viability of PDCs in SPC-A1 cells compared with that of polyethylenimine (PEI)/DNA complexes at various N/P ratios (* P
    Figure Legend Snippet: Transfection efficiency of polycation lipid nanocarrier/DNA complexes (PDC) (N/P = 10) in human lung adenocarcinoma (SPC-A1) cells compared with those of Lipofectamine™ 2000/DNA complexes (LDC). ( A ) Transfection efficiency of plasmid pEGFP-N2 determined by flow cytometer; ( B ) relative luciferase activity in SPC-A1 cells treated with PDC in comparison with that of LDC; ( C ) the cell viability of PDCs in SPC-A1 cells compared with that of polyethylenimine (PEI)/DNA complexes at various N/P ratios (* P

    Techniques Used: Transfection, Plasmid Preparation, Flow Cytometry, Cytometry, Luciferase, Activity Assay

    Transfection of pEGFP-N2 in human lung adenocarcinoma (SPC-A1) cells mediated with Lipofectamine™ 2000/DNA complexes (LDC) and polycation lipid nanocarrier/DNA complexes (PDC) (N/P = 10). ( A ) Images of SPC-A1 cells transfected by LDC, scale bar = 100 μm; ( B ) images of SPC-A1 cells mediated by PDC (N/P = 10), scale bar = 100 μm; ( C ) fluorescence intensity of expressed green fluorescent protein in SPC-A1 cells mediated by different PDCs with various N/P ratios (** P
    Figure Legend Snippet: Transfection of pEGFP-N2 in human lung adenocarcinoma (SPC-A1) cells mediated with Lipofectamine™ 2000/DNA complexes (LDC) and polycation lipid nanocarrier/DNA complexes (PDC) (N/P = 10). ( A ) Images of SPC-A1 cells transfected by LDC, scale bar = 100 μm; ( B ) images of SPC-A1 cells mediated by PDC (N/P = 10), scale bar = 100 μm; ( C ) fluorescence intensity of expressed green fluorescent protein in SPC-A1 cells mediated by different PDCs with various N/P ratios (** P

    Techniques Used: Transfection, Fluorescence

    12) Product Images from "MicroRNA 21 Promotes Glioma Invasion by Targeting Matrix Metalloproteinase Regulators ▿MicroRNA 21 Promotes Glioma Invasion by Targeting Matrix Metalloproteinase Regulators ▿ †"

    Article Title: MicroRNA 21 Promotes Glioma Invasion by Targeting Matrix Metalloproteinase Regulators ▿MicroRNA 21 Promotes Glioma Invasion by Targeting Matrix Metalloproteinase Regulators ▿ †

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.00479-08

    Inhibition of miR-21 affects glioma cell motility. (A) Validation of RECK downregulation by siRNA. A172 cells were transfected with RECK siRNA or Lipofectamine only (mock). At 72 h following transfection, cells were harvested and subjected to Western
    Figure Legend Snippet: Inhibition of miR-21 affects glioma cell motility. (A) Validation of RECK downregulation by siRNA. A172 cells were transfected with RECK siRNA or Lipofectamine only (mock). At 72 h following transfection, cells were harvested and subjected to Western

    Techniques Used: Inhibition, Transfection, Western Blot

    13) Product Images from "Construction of the plasmid coding for the expression of the EGFP-M-IL-2(88Arg, 125Ala) fusion protein and the anti-tumor effects exerted by the fusion protein in HeLa-60 cells"

    Article Title: Construction of the plasmid coding for the expression of the EGFP-M-IL-2(88Arg, 125Ala) fusion protein and the anti-tumor effects exerted by the fusion protein in HeLa-60 cells

    Journal: Oncology Letters

    doi: 10.3892/ol.2015.3125

    The morphology of various HeLa cell treatment groups were observed at 24, 48 and 72 h subsequnet to transfection. (A) Cells transfected with pLEGFP-C1- M-IL -2( 88 Arg,  125 Ala). (B) pLEGFP-C1-transfected cells. (C) Lipofectamine 2000-transfected cells. (D)
    Figure Legend Snippet: The morphology of various HeLa cell treatment groups were observed at 24, 48 and 72 h subsequnet to transfection. (A) Cells transfected with pLEGFP-C1- M-IL -2( 88 Arg, 125 Ala). (B) pLEGFP-C1-transfected cells. (C) Lipofectamine 2000-transfected cells. (D)

    Techniques Used: Transfection

    14) Product Images from "Short interfering RNA-mediated silencing of actin-related protein 2/3 complex subunit 4 inhibits the migration of SW620 human colorectal cancer cells"

    Article Title: Short interfering RNA-mediated silencing of actin-related protein 2/3 complex subunit 4 inhibits the migration of SW620 human colorectal cancer cells

    Journal: Oncology Letters

    doi: 10.3892/ol.2017.7642

    Growth curves for cells subsequent to actin-related protein 2/3 complex subunit 4 siRNA transfection. (A) OD 450 value of each group. (B) Mean absorbance of each group. The relative proliferation rates at 0, 1, 2, 3 and 4 days subsequent to transfection in SW620 cells show no significant differences between the siRNA group and NC group (ns vs. NC). OD, optical density; siRNA, small interfering target RNA; NC, nonspecific siRNA; Lipo, Lipofectamine-only group; Null, control group; ns, not significant.
    Figure Legend Snippet: Growth curves for cells subsequent to actin-related protein 2/3 complex subunit 4 siRNA transfection. (A) OD 450 value of each group. (B) Mean absorbance of each group. The relative proliferation rates at 0, 1, 2, 3 and 4 days subsequent to transfection in SW620 cells show no significant differences between the siRNA group and NC group (ns vs. NC). OD, optical density; siRNA, small interfering target RNA; NC, nonspecific siRNA; Lipo, Lipofectamine-only group; Null, control group; ns, not significant.

    Techniques Used: Transfection

    Cell cycle distribution analysis by flow cytometry. (A) siRNA group; (B) NC group; (C) Lipo group and (D) Null group 24 h after transfection of SW620 cells. Differences in the G0/G1, S and G2/M phase distribution were not statistically significant between the siRNA group and NC group (ns vs. NC). siRNA, small interfering target RNA; Null, control group; NC, nonspecific siRNA; Lipo, Lipofectamine-only group; ns, not significant.
    Figure Legend Snippet: Cell cycle distribution analysis by flow cytometry. (A) siRNA group; (B) NC group; (C) Lipo group and (D) Null group 24 h after transfection of SW620 cells. Differences in the G0/G1, S and G2/M phase distribution were not statistically significant between the siRNA group and NC group (ns vs. NC). siRNA, small interfering target RNA; Null, control group; NC, nonspecific siRNA; Lipo, Lipofectamine-only group; ns, not significant.

    Techniques Used: Flow Cytometry, Cytometry, Transfection

    15) Product Images from "The natural polyphenol curcumin induces apoptosis by suppressing STAT3 signaling in esophageal squamous cell carcinoma"

    Article Title: The natural polyphenol curcumin induces apoptosis by suppressing STAT3 signaling in esophageal squamous cell carcinoma

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    doi: 10.1186/s13046-018-0959-0

    Curcumin inhibited STAT3 activation in ESCC cells.  a  TE13 cells were transfected with vector or STAT3-luc by Lipofectamine ®  2000 for 24 h. Cells were then treated with 0, 8 or 16 μM curcumin for 12 h. Luciferase activity was measured followed by stimulation with IL-6 or vehicle control for 20 min.  b  EC9706 and TE13 cells were treated with different concentrations of curcumin for 24 h. Then, cells were prepared for immunoblotting against p-STAT3, STAT3 and GAPDH.  c  EC9706 and TE13 cells were treated with 16 μM curcumin for indicated time, followed by immunoblotting against p-STAT3, STAT3 and β-actin.  d  Following starved overnight in serum-free medium, EC9706 and TE13 cells were incubated with 0, 4, 8 or 16 μM curcumin for 4 h, then stimulated with IL-6 (50 ng/ml) for 20 min. The cells were lysed and analyzed by immunoblotting against p-STAT3, STAT3 and GAPDH.  e  mRNA levels of Cyclin D1, MCL1 and IL-6 were measured by qRT-PCR in TE13 cells treated with 0, 4, 8 or 16 μM curcumin overnight. * p
    Figure Legend Snippet: Curcumin inhibited STAT3 activation in ESCC cells. a TE13 cells were transfected with vector or STAT3-luc by Lipofectamine ® 2000 for 24 h. Cells were then treated with 0, 8 or 16 μM curcumin for 12 h. Luciferase activity was measured followed by stimulation with IL-6 or vehicle control for 20 min. b EC9706 and TE13 cells were treated with different concentrations of curcumin for 24 h. Then, cells were prepared for immunoblotting against p-STAT3, STAT3 and GAPDH. c EC9706 and TE13 cells were treated with 16 μM curcumin for indicated time, followed by immunoblotting against p-STAT3, STAT3 and β-actin. d Following starved overnight in serum-free medium, EC9706 and TE13 cells were incubated with 0, 4, 8 or 16 μM curcumin for 4 h, then stimulated with IL-6 (50 ng/ml) for 20 min. The cells were lysed and analyzed by immunoblotting against p-STAT3, STAT3 and GAPDH. e mRNA levels of Cyclin D1, MCL1 and IL-6 were measured by qRT-PCR in TE13 cells treated with 0, 4, 8 or 16 μM curcumin overnight. * p

    Techniques Used: Activation Assay, Transfection, Plasmid Preparation, Luciferase, Activity Assay, Incubation, Quantitative RT-PCR

    16) Product Images from "Inhibitory Effect and Mechanism of Arctium lappa Extract on NLRP3 Inflammasome Activation"

    Article Title: Inhibitory Effect and Mechanism of Arctium lappa Extract on NLRP3 Inflammasome Activation

    Journal: Evidence-based Complementary and Alternative Medicine : eCAM

    doi: 10.1155/2018/6346734

    Inhibitory effects of ALE on NLRP3 inflammasome mediated IL-1 β  secretion  in vitro  and  in vivo . (a) BMDMs were treated with the indicated concentration of ALE for 6 h. Cell viability was measured by MTT assay. LPS-primed BMDMs were pretreated with ALE or zVAD at an indicated concentration for 1 h and then stimulated with ATP (b, e, and g); nigericin (Nig.) (c, f, and h) for 1 h; and silica crystals (silica) (d and i) for 3 h. IL-1 β  (b–d) and TNF- α  (f) concentrations in the culture supernatant were measured by ELISA. (e) Cytotoxicity of ALE was measured with LDH release in the culture supernatants. (g~i) Culture supernatants (S/N) and cell lysates from BMDMs pretreated with or without sample and primed with LPS and indicated stimulators were analyzed by immunoblotting. LPS-primed BMDMs were treated with ALE or zVAD at an indicated concentration for 1 h, and flagellin (Fla) (j) or poly(dA:dT) (k) was delivered into cytosol through lipofectamine 2000. Culture supernatants were analyzed by immunoblotting. (l and m) C57BL/6 mice were given intraperitoneal injections of ALE or a NLRP3 specific inhibitor, MCC950, 2 h and 12 h prior to LPS injection (20 mg/kg). Blood samples were collected 2 h after the LPS challenge and the concentrations of IL-1 β  (l) and TNF- α  (m) in plasma were measured by ELISA. The data represent the mean ± SEM of three independent experiments performed in triplicate.  ∗ p
    Figure Legend Snippet: Inhibitory effects of ALE on NLRP3 inflammasome mediated IL-1 β secretion in vitro and in vivo . (a) BMDMs were treated with the indicated concentration of ALE for 6 h. Cell viability was measured by MTT assay. LPS-primed BMDMs were pretreated with ALE or zVAD at an indicated concentration for 1 h and then stimulated with ATP (b, e, and g); nigericin (Nig.) (c, f, and h) for 1 h; and silica crystals (silica) (d and i) for 3 h. IL-1 β (b–d) and TNF- α (f) concentrations in the culture supernatant were measured by ELISA. (e) Cytotoxicity of ALE was measured with LDH release in the culture supernatants. (g~i) Culture supernatants (S/N) and cell lysates from BMDMs pretreated with or without sample and primed with LPS and indicated stimulators were analyzed by immunoblotting. LPS-primed BMDMs were treated with ALE or zVAD at an indicated concentration for 1 h, and flagellin (Fla) (j) or poly(dA:dT) (k) was delivered into cytosol through lipofectamine 2000. Culture supernatants were analyzed by immunoblotting. (l and m) C57BL/6 mice were given intraperitoneal injections of ALE or a NLRP3 specific inhibitor, MCC950, 2 h and 12 h prior to LPS injection (20 mg/kg). Blood samples were collected 2 h after the LPS challenge and the concentrations of IL-1 β (l) and TNF- α (m) in plasma were measured by ELISA. The data represent the mean ± SEM of three independent experiments performed in triplicate. ∗ p

    Techniques Used: In Vitro, In Vivo, Concentration Assay, MTT Assay, Enzyme-linked Immunosorbent Assay, Mouse Assay, Injection

    17) Product Images from "Placental trophoblast debris mediated feto-maternal signalling via small RNA delivery: implications for preeclampsia"

    Article Title: Placental trophoblast debris mediated feto-maternal signalling via small RNA delivery: implications for preeclampsia

    Journal: Scientific Reports

    doi: 10.1038/s41598-017-14180-8

    Visualizing delivery of fluorescent siRNA to placental explants, trophoblast debris and transfer to endothelial cells by trophoblast debris. Normotensive term placental explants (n = 3) were cultured with ( A , B ) or without (bottom panel: D,E) the presence of 25 pmol of cy3 labelled siRNA (red) and lipofectamine mixture for 17 hours. Five µm sections were cut from the explants ( A , D ) and smears of trophoblast debris from the explants ( B , E ) were co-stained with DAPI nuclear marker (blue) and visualised by confocal microscopy. HMEC-1 cells were exposed to trophoblast debris from cy3-siRNA containing ( C , red) or untreated trophoblast debris ( F ) for 21 hours before co-staining with Cell Tracker Green (CMFDA, green) and DAPI (blue). The main panel of image C shows the projection image of the z-stack. The bottom panel shows the Z-projection of X-Z axis, while the panel on the right shows the Z-projection of Y-Z axis. Scale bars represent 50 μm in all images.
    Figure Legend Snippet: Visualizing delivery of fluorescent siRNA to placental explants, trophoblast debris and transfer to endothelial cells by trophoblast debris. Normotensive term placental explants (n = 3) were cultured with ( A , B ) or without (bottom panel: D,E) the presence of 25 pmol of cy3 labelled siRNA (red) and lipofectamine mixture for 17 hours. Five µm sections were cut from the explants ( A , D ) and smears of trophoblast debris from the explants ( B , E ) were co-stained with DAPI nuclear marker (blue) and visualised by confocal microscopy. HMEC-1 cells were exposed to trophoblast debris from cy3-siRNA containing ( C , red) or untreated trophoblast debris ( F ) for 21 hours before co-staining with Cell Tracker Green (CMFDA, green) and DAPI (blue). The main panel of image C shows the projection image of the z-stack. The bottom panel shows the Z-projection of X-Z axis, while the panel on the right shows the Z-projection of Y-Z axis. Scale bars represent 50 μm in all images.

    Techniques Used: Cell Culture, Staining, Marker, Confocal Microscopy

    18) Product Images from "The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation"

    Article Title: The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation

    Journal: Cells

    doi: 10.3390/cells9040825

    HspBP1 knockdown impairs SG formation. HeLa cells were incubated for 4 days with the transfection agent lipofectamine, mock-treated, transfected with a control plasmid (scrambled) or constructs targeting two different regions of the HspBP1 transcript (shRNA1 and shRNA2). ( A ) Crude cell extracts were probed for HspBP1; actin was used as loading control. Quantitative Western blotting revealed a significant depletion of HspBP1 for each of the sh-constructs. All lanes were on the same blot. Results are shown as average +SEM. Significant differences were identified with One-Way ANOVA combined with Bonferroni posthoc analysis; * p
    Figure Legend Snippet: HspBP1 knockdown impairs SG formation. HeLa cells were incubated for 4 days with the transfection agent lipofectamine, mock-treated, transfected with a control plasmid (scrambled) or constructs targeting two different regions of the HspBP1 transcript (shRNA1 and shRNA2). ( A ) Crude cell extracts were probed for HspBP1; actin was used as loading control. Quantitative Western blotting revealed a significant depletion of HspBP1 for each of the sh-constructs. All lanes were on the same blot. Results are shown as average +SEM. Significant differences were identified with One-Way ANOVA combined with Bonferroni posthoc analysis; * p

    Techniques Used: Incubation, Transfection, Plasmid Preparation, Construct, Western Blot

    Production of HspBP1 full length protein and deletion mutants. ( A ) Organization of the HspBP1 protein. HspBP1 domains and the regions interacting with hsc70 are shown for the full length protein [ 16 ]. The organization of deletion mutants is also depicted. ( B ) Western blot analysis of full length and mutant HspBP1. Crude extracts were prepared for transfected HeLa cells and probed with antibodies against HspBP1 or hsp72. Actin was used as loading control. Lipo, lipofectamine; ctl, control plasmid; full length HspBP1, ΔM (deletion of residues 154–195), ΔMC (deletion of residues 154–195 and 314–359) ΔC (deletion of residues 314–359). All lanes were on the same blot.
    Figure Legend Snippet: Production of HspBP1 full length protein and deletion mutants. ( A ) Organization of the HspBP1 protein. HspBP1 domains and the regions interacting with hsc70 are shown for the full length protein [ 16 ]. The organization of deletion mutants is also depicted. ( B ) Western blot analysis of full length and mutant HspBP1. Crude extracts were prepared for transfected HeLa cells and probed with antibodies against HspBP1 or hsp72. Actin was used as loading control. Lipo, lipofectamine; ctl, control plasmid; full length HspBP1, ΔM (deletion of residues 154–195), ΔMC (deletion of residues 154–195 and 314–359) ΔC (deletion of residues 314–359). All lanes were on the same blot.

    Techniques Used: Western Blot, Mutagenesis, Transfection, Plasmid Preparation

    19) Product Images from "A non-covalent peptide-based carrier for in vivo delivery of DNA mimics"

    Article Title: A non-covalent peptide-based carrier for in vivo delivery of DNA mimics

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkm053

    Pep-3-mediated delivery of an antisense cyclin B1 HypNA- p PNA in primary and suspension cell lines. ( A ) Pep-3-mediated delivery of antisense cyclin B1 HypNA- p PNA was evaluated on different cell lines. Here, 100 nM of antisense HypNA- p PNA was complexed with Pep-3 at a molar ratio 1:20, then evaluated on Jurkat T (lane 3), HUVEC (lane 4), PC3 (lane 5) and MCF7 (lane 6). Also, 100 nM of antisense HypNA- p PNA was complexed with Pep-2 at the same ratio and evaluated on Jurkat T (lane 2). The level of endogenous cyclin B1 in untreated cells is reported in lane 1. ( B/C ) Comparison of Pep-3 strategy to other delivery methods. Increasing concentrations from 5 nM to 1 µM of HypNA- p PNA Cyc-B1a were associated with Pep-3, Pep-2 or MPG at a molar ratio 1:20 or with Lipofectamine, then overlaid onto cultured HUVEC (panel B) and Jurkat (panel C) cells. Cyclin B1 protein levels were analysed by western blotting after 30 h and antisense effect was normalized. ( D/E ): Toxicity of the different delivery methods: HUVEC (panel E) and Jurkat (panel F) cells, were incubated with increasing concentrations of Cyc-B1 mm HypNA- p PNA complexed with Pep-3, Pep-2, MPG and Lipofectamine. Cell viability was evaluated by MTT assay after 24 h.
    Figure Legend Snippet: Pep-3-mediated delivery of an antisense cyclin B1 HypNA- p PNA in primary and suspension cell lines. ( A ) Pep-3-mediated delivery of antisense cyclin B1 HypNA- p PNA was evaluated on different cell lines. Here, 100 nM of antisense HypNA- p PNA was complexed with Pep-3 at a molar ratio 1:20, then evaluated on Jurkat T (lane 3), HUVEC (lane 4), PC3 (lane 5) and MCF7 (lane 6). Also, 100 nM of antisense HypNA- p PNA was complexed with Pep-2 at the same ratio and evaluated on Jurkat T (lane 2). The level of endogenous cyclin B1 in untreated cells is reported in lane 1. ( B/C ) Comparison of Pep-3 strategy to other delivery methods. Increasing concentrations from 5 nM to 1 µM of HypNA- p PNA Cyc-B1a were associated with Pep-3, Pep-2 or MPG at a molar ratio 1:20 or with Lipofectamine, then overlaid onto cultured HUVEC (panel B) and Jurkat (panel C) cells. Cyclin B1 protein levels were analysed by western blotting after 30 h and antisense effect was normalized. ( D/E ): Toxicity of the different delivery methods: HUVEC (panel E) and Jurkat (panel F) cells, were incubated with increasing concentrations of Cyc-B1 mm HypNA- p PNA complexed with Pep-3, Pep-2, MPG and Lipofectamine. Cell viability was evaluated by MTT assay after 24 h.

    Techniques Used: Cell Culture, Western Blot, Incubation, MTT Assay

    20) Product Images from "A shear stress responsive gene product PP1201 protects against Fas-mediated apoptosis by reducing Fas expression on the cell surface"

    Article Title: A shear stress responsive gene product PP1201 protects against Fas-mediated apoptosis by reducing Fas expression on the cell surface

    Journal: Apoptosis : an international journal on programmed cell death

    doi: 10.1007/s10495-010-0556-y

    Silencing of endogenous PP1201 does not influence Cell surface Fas expression but overexpressed PP1201 does. a Endogenous expression of PP1201 goes down up to 72 h upon silencing in HT1080 cells. 50 nM of ON-TARGET plus SMART pool (Thermo scientific) specific to human PP1201 was transfected using Lipofectamine RNAiMAX (Invitrogen). As a control 50 nM Non-targeting pool was used. After transfections lysates were prepared of transfected cells for transfection period of 24 h, 48 h and at 72 h and subjected to immunoblot with anti-PP1201 antibody. Blotting with β-actin shows loading control. 30 μg of protein was loaded in each lane. b Flow cytometry analysis shows cell surface level of Fas in HT1080 after silencing of endogenous PP1201 at 48 h. Overlay histogram represents one of the triplicate and inset shows average of mean fluorescence intensities (±SD) of triplicate values for Fas staining. c PP1210 expressing HT1080 cells were transfected with the indicated siRNAs and 30 μg of cellular lysates was probed with the indicated proteins. d Flow cytometry analysis exhibits surface levels of Fas after silencing to various levels of overexpressed PP1201 in HT1080 stable cell line. Overlay histogram represents one of the triplicates and inset shows average of mean fluorescence intensities (±SD) of triplicate values for Fas staining. e Overexpression of PP1201 in AoSMC cells down regulates the cell surface expression of Fas. AoSMC cells were transfected with 1 μg eGFP either with 5 μg vector (control) or with 5 μg PP1201 expression plasmid and after 48 h cells were stained with Fas antibody conjugated with APC and double positive cells were analyzed by FACS. Double positive populations were selected and Fas staining was analyzed by overlay histogram plot and by comparing mean fluorescence intensities. Overlay histogram represents one of the triplicates and inset shows average of mean fluorescence intensities (±SD) of triplicate values for Fas staining
    Figure Legend Snippet: Silencing of endogenous PP1201 does not influence Cell surface Fas expression but overexpressed PP1201 does. a Endogenous expression of PP1201 goes down up to 72 h upon silencing in HT1080 cells. 50 nM of ON-TARGET plus SMART pool (Thermo scientific) specific to human PP1201 was transfected using Lipofectamine RNAiMAX (Invitrogen). As a control 50 nM Non-targeting pool was used. After transfections lysates were prepared of transfected cells for transfection period of 24 h, 48 h and at 72 h and subjected to immunoblot with anti-PP1201 antibody. Blotting with β-actin shows loading control. 30 μg of protein was loaded in each lane. b Flow cytometry analysis shows cell surface level of Fas in HT1080 after silencing of endogenous PP1201 at 48 h. Overlay histogram represents one of the triplicate and inset shows average of mean fluorescence intensities (±SD) of triplicate values for Fas staining. c PP1210 expressing HT1080 cells were transfected with the indicated siRNAs and 30 μg of cellular lysates was probed with the indicated proteins. d Flow cytometry analysis exhibits surface levels of Fas after silencing to various levels of overexpressed PP1201 in HT1080 stable cell line. Overlay histogram represents one of the triplicates and inset shows average of mean fluorescence intensities (±SD) of triplicate values for Fas staining. e Overexpression of PP1201 in AoSMC cells down regulates the cell surface expression of Fas. AoSMC cells were transfected with 1 μg eGFP either with 5 μg vector (control) or with 5 μg PP1201 expression plasmid and after 48 h cells were stained with Fas antibody conjugated with APC and double positive cells were analyzed by FACS. Double positive populations were selected and Fas staining was analyzed by overlay histogram plot and by comparing mean fluorescence intensities. Overlay histogram represents one of the triplicates and inset shows average of mean fluorescence intensities (±SD) of triplicate values for Fas staining

    Techniques Used: Expressing, Transfection, Flow Cytometry, Fluorescence, Staining, Stable Transfection, Over Expression, Plasmid Preparation, FACS

    21) Product Images from "Detection of mRNA in living cells by double-stranded locked nucleic acid probes"

    Article Title: Detection of mRNA in living cells by double-stranded locked nucleic acid probes

    Journal: The Analyst

    doi: 10.1039/c3an00722g

    Delivery of dsLNA probes to cells. Transfection efficiency with (a) different transfection reagents and (b) different amounts of Lipofectamine 2000 per 1.0 µg dsLNA probe. (c) Photothermal delivery of dsLNA probes targeting β-actin mRNA
    Figure Legend Snippet: Delivery of dsLNA probes to cells. Transfection efficiency with (a) different transfection reagents and (b) different amounts of Lipofectamine 2000 per 1.0 µg dsLNA probe. (c) Photothermal delivery of dsLNA probes targeting β-actin mRNA

    Techniques Used: Transfection

    22) Product Images from "Massive endocytosis triggered by surface membrane palmitoylation under mitochondrial control in BHK fibroblasts"

    Article Title: Massive endocytosis triggered by surface membrane palmitoylation under mitochondrial control in BHK fibroblasts

    Journal: eLife

    doi: 10.7554/eLife.01293

    The acyl tranferase, DHHC5, is required for MEND in BHK cells. ( A ) DHHC5 knockdown by siRNA to ZDHHC5. BHK cells were transfected with control siRNA or ZDHHC5-specific siRNA at indicated concentrations using Lipofectamine 2000. 72 hrs after transfection, cells were harvested and cell lysates were processed for Western blot analysis using 20 µg protein per lane. A human non-small lung carcinoma cell line, H1299, was used as a positive control for anti-ZDHHC5 antibody. ( B ) MEND in BHK cells amounts to 75% of the cell surface after growing cells with staurosporin (0.1 μM), and DHHC5 siRNA decreases MEND by 63% (p
    Figure Legend Snippet: The acyl tranferase, DHHC5, is required for MEND in BHK cells. ( A ) DHHC5 knockdown by siRNA to ZDHHC5. BHK cells were transfected with control siRNA or ZDHHC5-specific siRNA at indicated concentrations using Lipofectamine 2000. 72 hrs after transfection, cells were harvested and cell lysates were processed for Western blot analysis using 20 µg protein per lane. A human non-small lung carcinoma cell line, H1299, was used as a positive control for anti-ZDHHC5 antibody. ( B ) MEND in BHK cells amounts to 75% of the cell surface after growing cells with staurosporin (0.1 μM), and DHHC5 siRNA decreases MEND by 63% (p

    Techniques Used: Transfection, Western Blot, Positive Control

    23) Product Images from "Mirk/dyrk1B kinase is upregulated following inhibition of mTOR"

    Article Title: Mirk/dyrk1B kinase is upregulated following inhibition of mTOR

    Journal: Carcinogenesis

    doi: 10.1093/carcin/bgu058

    Mirk mRNA levels increased by either depletion or inhibition of mTOR and decreased following CREB depletion. ( A ) Northern analysis of Mirk mRNA levels. Left: Panc1 cells treated with increasing concentrations of WYE354 for 1 day before northern analysis (lanes 2–5). Panc1 cells were transfected (5 µl RNA interference (RNAi)/5 µl Lipofectamine 2000) for 1 day in complete medium with either RNAi FRAP1HSS103825 (T1) to mTOR/FRAP, RNAi C90 to CREB or a GC-matched negative control RNAi (Ct) then switched for 1 day serum-free DMEM to allow Mirk expression before analysis, lanes 6–8. Positive control for high Mirk expression, 1 day serum-free culture. Right: Panc1 cells were transfected either RNAi’s FRAP1HSS103825 (T1) or FRAP1HSS103827 (T3) to mTOR/FRAP, RNAi’s C90 or C91 to CREB or a GC-matched negative control RNAi (Ct), then switched for 1 day to serum-free DMEM to allow Mirk expression before analysis. One of the duplicate experiments with similar results. Below each northern blot is an ethidium bromide-stained total RNA panel showing 28s and 18s rRNAs. ( B ) Western blot of CREB and mTOR protein levels after Panc1 cell treatment with RNAi’s in panel A. C90 and C91 reduced CREB levels 60%, whereas T1 and T3 reduced mTOR levels 40% and 70%, respectively (Uni-Scan densitometry). ( C ) Graph showing increase in Mirk mRNA with increasing WYE354 concentration. ( D ) Summary of northern analysis of Mirk mRNA levels. Data on mRNA expression from panel A pooled, together with five samples treated with mixtures of RNA’s to CREB and to mTOR, mean ± SE shown. Dotted line indicates control level. The mean of the mTORsi and the mean of the CREBsi were statistically different, P = 0.022 by two-tailed t -test. CREBsi, small interfering RNA to CREB; FRAP, FKBP12-rapamycin complex-associated protein; PTEN, phosphatase and tensin homolog; mTORsi, small interfering RNA to mTOR.
    Figure Legend Snippet: Mirk mRNA levels increased by either depletion or inhibition of mTOR and decreased following CREB depletion. ( A ) Northern analysis of Mirk mRNA levels. Left: Panc1 cells treated with increasing concentrations of WYE354 for 1 day before northern analysis (lanes 2–5). Panc1 cells were transfected (5 µl RNA interference (RNAi)/5 µl Lipofectamine 2000) for 1 day in complete medium with either RNAi FRAP1HSS103825 (T1) to mTOR/FRAP, RNAi C90 to CREB or a GC-matched negative control RNAi (Ct) then switched for 1 day serum-free DMEM to allow Mirk expression before analysis, lanes 6–8. Positive control for high Mirk expression, 1 day serum-free culture. Right: Panc1 cells were transfected either RNAi’s FRAP1HSS103825 (T1) or FRAP1HSS103827 (T3) to mTOR/FRAP, RNAi’s C90 or C91 to CREB or a GC-matched negative control RNAi (Ct), then switched for 1 day to serum-free DMEM to allow Mirk expression before analysis. One of the duplicate experiments with similar results. Below each northern blot is an ethidium bromide-stained total RNA panel showing 28s and 18s rRNAs. ( B ) Western blot of CREB and mTOR protein levels after Panc1 cell treatment with RNAi’s in panel A. C90 and C91 reduced CREB levels 60%, whereas T1 and T3 reduced mTOR levels 40% and 70%, respectively (Uni-Scan densitometry). ( C ) Graph showing increase in Mirk mRNA with increasing WYE354 concentration. ( D ) Summary of northern analysis of Mirk mRNA levels. Data on mRNA expression from panel A pooled, together with five samples treated with mixtures of RNA’s to CREB and to mTOR, mean ± SE shown. Dotted line indicates control level. The mean of the mTORsi and the mean of the CREBsi were statistically different, P = 0.022 by two-tailed t -test. CREBsi, small interfering RNA to CREB; FRAP, FKBP12-rapamycin complex-associated protein; PTEN, phosphatase and tensin homolog; mTORsi, small interfering RNA to mTOR.

    Techniques Used: Inhibition, Northern Blot, Transfection, Negative Control, Expressing, Positive Control, Staining, Western Blot, Concentration Assay, Two Tailed Test, Small Interfering RNA

    Mirk promoter activated through CREB binding sites. ( A ) Dose-dependent activation of Mirk promoter–luciferase reporter by increasing amounts of transfected CREB. Panc1 cells were transfected with 0.5 µg pMp (Nhe1-EcoRv-luciferase 9657kb) Mirk promoter–reporter and 0.25 µg β-galactosidase reporter plus increasing amounts of the CREB expression plasmid. Cells were cultured overnight either in serum-free DMEM or serum-containing growth medium before lysis and assay. Data are mean ± SD ( n = 3). ( B ) Panc1 cells were transfected with 0.5 µg of either the 9600bp Nhe1-EcoRv-luciferase Mirk promoter–reporter containing two upstream putative CREB binding sites and one site in exon 4, the 5887bp NheI-Sma Mirk promoter–reporter containing two upstream putative CREB binding sites only or the 4830bp HincII-Sca1 Mirk promoter–reporter containing only the putative CRE binding site in exon 4. Cotransfected were 0.25 µg β-galactosidase reporter plus 0.5 µg of either the wild-type or S133A mutant CREB expression plasmid. Cells were cultured overnight in serum-free DMEM to allow Mirk expression before lysis and assay. Data are shown as mean ± SD ( n = 3). ( C ) C2C12 myoblasts in 12-well plates were transfected for 4h with 2 µl Lipofectamine Plus and the 0.25 µg β-galactosidase reporter, 0.5 µg of either the wild-type or S133A mutant CREB expression plasmid and 5 µg of the three Mp-luciferase constructs listed in panel B plus 2 mutant constructs. The cells were cultured overnight, in DMEM + 10% FBS, then washed and placed in differentiation medium (DMEM plus 2% horse serum) for 24h before assay. Data are shown as mean ± SD ( n = 3). Results are one of three similar experiments. The numbers above the bars indicate whether the construct has one or two upstream CREB sites/downstream site, either wild-type or mutant (m).
    Figure Legend Snippet: Mirk promoter activated through CREB binding sites. ( A ) Dose-dependent activation of Mirk promoter–luciferase reporter by increasing amounts of transfected CREB. Panc1 cells were transfected with 0.5 µg pMp (Nhe1-EcoRv-luciferase 9657kb) Mirk promoter–reporter and 0.25 µg β-galactosidase reporter plus increasing amounts of the CREB expression plasmid. Cells were cultured overnight either in serum-free DMEM or serum-containing growth medium before lysis and assay. Data are mean ± SD ( n = 3). ( B ) Panc1 cells were transfected with 0.5 µg of either the 9600bp Nhe1-EcoRv-luciferase Mirk promoter–reporter containing two upstream putative CREB binding sites and one site in exon 4, the 5887bp NheI-Sma Mirk promoter–reporter containing two upstream putative CREB binding sites only or the 4830bp HincII-Sca1 Mirk promoter–reporter containing only the putative CRE binding site in exon 4. Cotransfected were 0.25 µg β-galactosidase reporter plus 0.5 µg of either the wild-type or S133A mutant CREB expression plasmid. Cells were cultured overnight in serum-free DMEM to allow Mirk expression before lysis and assay. Data are shown as mean ± SD ( n = 3). ( C ) C2C12 myoblasts in 12-well plates were transfected for 4h with 2 µl Lipofectamine Plus and the 0.25 µg β-galactosidase reporter, 0.5 µg of either the wild-type or S133A mutant CREB expression plasmid and 5 µg of the three Mp-luciferase constructs listed in panel B plus 2 mutant constructs. The cells were cultured overnight, in DMEM + 10% FBS, then washed and placed in differentiation medium (DMEM plus 2% horse serum) for 24h before assay. Data are shown as mean ± SD ( n = 3). Results are one of three similar experiments. The numbers above the bars indicate whether the construct has one or two upstream CREB sites/downstream site, either wild-type or mutant (m).

    Techniques Used: Binding Assay, Activation Assay, Luciferase, Transfection, Expressing, Plasmid Preparation, Cell Culture, Lysis, Mutagenesis, Construct

    24) Product Images from "Tumor Necrosis Factor-Like Weak Inducer of Apoptosis Activates Type I Interferon Signals in Lupus Nephritis"

    Article Title: Tumor Necrosis Factor-Like Weak Inducer of Apoptosis Activates Type I Interferon Signals in Lupus Nephritis

    Journal: BioMed Research International

    doi: 10.1155/2017/4927376

    TWEAK mRNA expression was suppressed by TWEAK-siRNA-lipofectamine 2000 in PBMCs. Total RNA from PBMCs obtained from LN patients ( n  = 12) was extracted for quantitative real-time PCR. The ratio of TWEAK/ β -actin mRNA levels was calculated. The ratio of mRNA levels in cells treated with normal saline was designated as one. The error bars represent standard deviations. Control, PBMCs treated with normal saline; TWEAK-siRNA, PBMCs treated with TWEAK-siRNA-lipofectamine 2000; control-siRNA, PBMCs treated with control-siRNA-lipofectamine 2000.  † p
    Figure Legend Snippet: TWEAK mRNA expression was suppressed by TWEAK-siRNA-lipofectamine 2000 in PBMCs. Total RNA from PBMCs obtained from LN patients ( n = 12) was extracted for quantitative real-time PCR. The ratio of TWEAK/ β -actin mRNA levels was calculated. The ratio of mRNA levels in cells treated with normal saline was designated as one. The error bars represent standard deviations. Control, PBMCs treated with normal saline; TWEAK-siRNA, PBMCs treated with TWEAK-siRNA-lipofectamine 2000; control-siRNA, PBMCs treated with control-siRNA-lipofectamine 2000. † p

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction

    TWEAK upregulated expression of type I IFN-inducible genes in PBMCs. Total RNA from PBMCs obtained from LN patients ( n  = 12) was extracted for quantitative real-time PCR. The ratio of target gene/ β -actin mRNA levels was calculated. The ratio of mRNA levels in cells treated with normal saline was designated as one. The error bars represent standard deviations. Control, PBMCs treated with normal saline; TWEAK-siRNA, PBMCs treated with TWEAK-siRNA-lipofectamine 2000; control-siRNA, PBMCs treated with control-siRNA-lipofectamine 2000. rhTWEAK, PBMCs treated with rhTWEAK.  † p
    Figure Legend Snippet: TWEAK upregulated expression of type I IFN-inducible genes in PBMCs. Total RNA from PBMCs obtained from LN patients ( n = 12) was extracted for quantitative real-time PCR. The ratio of target gene/ β -actin mRNA levels was calculated. The ratio of mRNA levels in cells treated with normal saline was designated as one. The error bars represent standard deviations. Control, PBMCs treated with normal saline; TWEAK-siRNA, PBMCs treated with TWEAK-siRNA-lipofectamine 2000; control-siRNA, PBMCs treated with control-siRNA-lipofectamine 2000. rhTWEAK, PBMCs treated with rhTWEAK. † p

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction

    25) Product Images from "HEG1 is a novel mucin-like membrane protein that serves as a diagnostic and therapeutic target for malignant mesothelioma"

    Article Title: HEG1 is a novel mucin-like membrane protein that serves as a diagnostic and therapeutic target for malignant mesothelioma

    Journal: Scientific Reports

    doi: 10.1038/srep45768

    Suppression of proliferation of mesothelioma cells by HEG1 siRNA. Values are means ± S.D. of tetraplicate determinations. ( a ) Time-course of cell growth of ACC-MESO-4 treated with HEG1 siRNA. siRNA was transfected with Lipofectamine 2000. HEG1 siRNA mix 1, a mixture of H1097 and H2674 (1:1); HEG1 siRNA mix 2, sc-78365 (Santa Cruz Biotechnology); Control siRNA, MISSION siRNA Universal Negative Control (SIC-001). Similar results were obtained in 2 independent experiments. ( b ) Suppression of cell proliferation of ACC-MESO-4 with several HEG1 siRNAs. The cells were cultured for 72 h with siRNA and Lipofectamine 2000. Control siRNA, MISSION siRNA Universal Negative Control. Similar results were obtained in 2 independent experiments. ( c ) Suppression of cell proliferation with HEG1 siRNA on MPM cell lines. The cells were treated for 72 h with siRNA and Lipofectamine RNAiMAX. HEG1 expression after 72 h was shown in western blotting using mAb SKM9-2 at the bottom of the figure. Differences between mean values were analyzed by Student’s  t -test.  P -values for tests of ACC-MESO-4 and NCI-H2452 were
    Figure Legend Snippet: Suppression of proliferation of mesothelioma cells by HEG1 siRNA. Values are means ± S.D. of tetraplicate determinations. ( a ) Time-course of cell growth of ACC-MESO-4 treated with HEG1 siRNA. siRNA was transfected with Lipofectamine 2000. HEG1 siRNA mix 1, a mixture of H1097 and H2674 (1:1); HEG1 siRNA mix 2, sc-78365 (Santa Cruz Biotechnology); Control siRNA, MISSION siRNA Universal Negative Control (SIC-001). Similar results were obtained in 2 independent experiments. ( b ) Suppression of cell proliferation of ACC-MESO-4 with several HEG1 siRNAs. The cells were cultured for 72 h with siRNA and Lipofectamine 2000. Control siRNA, MISSION siRNA Universal Negative Control. Similar results were obtained in 2 independent experiments. ( c ) Suppression of cell proliferation with HEG1 siRNA on MPM cell lines. The cells were treated for 72 h with siRNA and Lipofectamine RNAiMAX. HEG1 expression after 72 h was shown in western blotting using mAb SKM9-2 at the bottom of the figure. Differences between mean values were analyzed by Student’s t -test. P -values for tests of ACC-MESO-4 and NCI-H2452 were

    Techniques Used: Transfection, Negative Control, Cell Culture, Expressing, Western Blot

    26) Product Images from "Dual-targeting nanoparticles with excellent gene transfection efficiency for gene therapy of peritoneal metastasis of colorectal cancer"

    Article Title: Dual-targeting nanoparticles with excellent gene transfection efficiency for gene therapy of peritoneal metastasis of colorectal cancer

    Journal: Oncotarget

    doi: 10.18632/oncotarget.21159

    Comparison of the transfection efficiency of PF 33 /pGFP (PF 33 ), RRPHC/pGFP (RRPHC) and Lipofectamine 3000/pGFP (Lipofectamine 3000) in medium containing 0%~30% serum in SW 480 cell ( A ) Images taken by fluorescence microscope. ( B , C ) Quantitative analysis of positive GFP cells (%) and Mean Fluorescence Intensity (MFI) by flow cytometry.
    Figure Legend Snippet: Comparison of the transfection efficiency of PF 33 /pGFP (PF 33 ), RRPHC/pGFP (RRPHC) and Lipofectamine 3000/pGFP (Lipofectamine 3000) in medium containing 0%~30% serum in SW 480 cell ( A ) Images taken by fluorescence microscope. ( B , C ) Quantitative analysis of positive GFP cells (%) and Mean Fluorescence Intensity (MFI) by flow cytometry.

    Techniques Used: Transfection, Fluorescence, Microscopy, Flow Cytometry, Cytometry

    Comparison of the transfection efficiency of PF 33 /pGFP (PF 33 ), RRPHC/pGFP (RRPHC) and Lipofectamine 2000/pGFP (Lipofectamine 2000) in medium containing 10%~30% serum in SW 480 cell ( A ) Images taken by fluorescence microscope. ( B ,  C ) Quantitative analysis of positive GFP cells (%) and Mean Fluorescence Intensity (MFI) by flow cytometry.
    Figure Legend Snippet: Comparison of the transfection efficiency of PF 33 /pGFP (PF 33 ), RRPHC/pGFP (RRPHC) and Lipofectamine 2000/pGFP (Lipofectamine 2000) in medium containing 10%~30% serum in SW 480 cell ( A ) Images taken by fluorescence microscope. ( B , C ) Quantitative analysis of positive GFP cells (%) and Mean Fluorescence Intensity (MFI) by flow cytometry.

    Techniques Used: Transfection, Fluorescence, Microscopy, Flow Cytometry, Cytometry

    27) Product Images from "A Human Periodontal Ligament Fibroblast Cell Line as a New Model to Study Periodontal Stress"

    Article Title: A Human Periodontal Ligament Fibroblast Cell Line as a New Model to Study Periodontal Stress

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms21217961

    Transfection efficiency of PDL-hTERT cells. Cells were transfected with PEI, SuperFect (SF) and Lipofectamine 2000 (LF) in low and high amounts. ( a ) Representative images of GFP-positive transfected cells (green) and DAPI-stained nuclei of all cells (blue). Scale bar: 100 µm. ( b ) Quantification of transfection efficiency. GFP-positive transfected cells per field of view were set in relation to the number of DAPI-positive cell nuclei per field of view and are presented as % ±SEM. Untransfected cells served as control (ctr). ( c ) Cell viability was calculated by counting DAPI-positive nuclei per field of view and set in relation to DAPI-positive nuclei per field of view of untransfected cells (ctr) and are presented as % ±SEM. n = 3 biological replicates performed as triplicates with three analyzed photographs per singlet. * 0.01 ≤ p
    Figure Legend Snippet: Transfection efficiency of PDL-hTERT cells. Cells were transfected with PEI, SuperFect (SF) and Lipofectamine 2000 (LF) in low and high amounts. ( a ) Representative images of GFP-positive transfected cells (green) and DAPI-stained nuclei of all cells (blue). Scale bar: 100 µm. ( b ) Quantification of transfection efficiency. GFP-positive transfected cells per field of view were set in relation to the number of DAPI-positive cell nuclei per field of view and are presented as % ±SEM. Untransfected cells served as control (ctr). ( c ) Cell viability was calculated by counting DAPI-positive nuclei per field of view and set in relation to DAPI-positive nuclei per field of view of untransfected cells (ctr) and are presented as % ±SEM. n = 3 biological replicates performed as triplicates with three analyzed photographs per singlet. * 0.01 ≤ p

    Techniques Used: Transfection, Staining

    28) Product Images from "Hydrophobic Oxime Ethers: A Versatile Class of pDNA and siRNA Transfection Lipids"

    Article Title: Hydrophobic Oxime Ethers: A Versatile Class of pDNA and siRNA Transfection Lipids

    Journal: ChemMedChem

    doi: 10.1002/cmdc.201100259

    Transfection of ( A ) MCF-7 cells and ( B ) H1792 cells using lipoplexes formulated at different N:P charge ratios. Results are expressed as total relative light units (RLU). Transfections were performed in 24-well tissue culture plates using 0.025 μg luciferase reporter construct (pCMVLuc) per well with 18h transfection time. Each data point reflects the mean value of three separate transfections. Error bars show the standard deviation from the mean. LFT = Lipofectamine 2000, jetP = jetPRIME.
    Figure Legend Snippet: Transfection of ( A ) MCF-7 cells and ( B ) H1792 cells using lipoplexes formulated at different N:P charge ratios. Results are expressed as total relative light units (RLU). Transfections were performed in 24-well tissue culture plates using 0.025 μg luciferase reporter construct (pCMVLuc) per well with 18h transfection time. Each data point reflects the mean value of three separate transfections. Error bars show the standard deviation from the mean. LFT = Lipofectamine 2000, jetP = jetPRIME.

    Techniques Used: Transfection, Luciferase, Construct, Standard Deviation

    Relative cytotoxicity MCF-7 (dark bars) and H1792 cells (light bars) were transfected using either lipofectamine 2000 (LFT), jetPrime (jP) or oxime ether lipoplexes at the specified N:P ratio (value in parenthesis). Transfections were performed using a 96-well format. Dead and live cells were counted 18 h post transfection using a trypan blue stain (n = 3 ).
    Figure Legend Snippet: Relative cytotoxicity MCF-7 (dark bars) and H1792 cells (light bars) were transfected using either lipofectamine 2000 (LFT), jetPrime (jP) or oxime ether lipoplexes at the specified N:P ratio (value in parenthesis). Transfections were performed using a 96-well format. Dead and live cells were counted 18 h post transfection using a trypan blue stain (n = 3 ).

    Techniques Used: Transfection, Staining

    GFP expression in PAR C10 cells ( A ) Cells were transfected (0.2 μg GFP reporter construct) with lipoplex formulations derived from lipids  2 – 5  (N:P 1.5) and excess LFT (positive control, Lipofectamine 2000, 1.6 μL); ( B ) Comparison of lipid  3  formulations at the indicated N:P ratio with LFT at the vendor’s suggested dose (0.5 μL). The transfection percentage of GFP expressing cells was analyzed by flow cytometry 48h post-transfection. Each data point reflects the mean ± SD from three independent experiments.
    Figure Legend Snippet: GFP expression in PAR C10 cells ( A ) Cells were transfected (0.2 μg GFP reporter construct) with lipoplex formulations derived from lipids 2 – 5 (N:P 1.5) and excess LFT (positive control, Lipofectamine 2000, 1.6 μL); ( B ) Comparison of lipid 3 formulations at the indicated N:P ratio with LFT at the vendor’s suggested dose (0.5 μL). The transfection percentage of GFP expressing cells was analyzed by flow cytometry 48h post-transfection. Each data point reflects the mean ± SD from three independent experiments.

    Techniques Used: Expressing, Transfection, Construct, Derivative Assay, Positive Control, Flow Cytometry, Cytometry

    Cell viability assay PAR C10 cells were transfected (N:P 1.5) with lipoplex formulations derived from lipids 2 – 5 and Lipofectamine 2000 (1.6 μL). After 48h, the cell viability was measured by calculating the % of alamar blue dye reduction. The data are shown as the mean ± SD of three independent experiments.
    Figure Legend Snippet: Cell viability assay PAR C10 cells were transfected (N:P 1.5) with lipoplex formulations derived from lipids 2 – 5 and Lipofectamine 2000 (1.6 μL). After 48h, the cell viability was measured by calculating the % of alamar blue dye reduction. The data are shown as the mean ± SD of three independent experiments.

    Techniques Used: Viability Assay, Transfection, Derivative Assay

    MCF-7 cells were transfected with a GFP expression plasmid (0.1 μg/well in a 24-well plate). Figures are presented as an overlay of green fluorescence (visualized by fluorescent microscopy after 20 hours of transfection) over the phase contrast image: ( A ) cells treated with pDNA (negative control); ( B ) cells transfected with lipoplexes derived from Lipofectamine 2000 (positive control, vendor’s recommended dose, 0.25 μL); ( C ) cells transfected with lipoplexes derived from lipid 3 at N:P=7.
    Figure Legend Snippet: MCF-7 cells were transfected with a GFP expression plasmid (0.1 μg/well in a 24-well plate). Figures are presented as an overlay of green fluorescence (visualized by fluorescent microscopy after 20 hours of transfection) over the phase contrast image: ( A ) cells treated with pDNA (negative control); ( B ) cells transfected with lipoplexes derived from Lipofectamine 2000 (positive control, vendor’s recommended dose, 0.25 μL); ( C ) cells transfected with lipoplexes derived from lipid 3 at N:P=7.

    Techniques Used: Transfection, Expressing, Plasmid Preparation, Fluorescence, Microscopy, Negative Control, Derivative Assay, Positive Control

    29) Product Images from "Transcriptional Regulation of the Novel Toll-like Receptor Tlr13 *"

    Article Title: Transcriptional Regulation of the Novel Toll-like Receptor Tlr13 *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M109.022541

    Suppression of the  Tlr13  gene promoter by LPS and PGN. A , Raw264.7 cells were transfected with  Tlr13  promoter p-341 plus  Renilla -TK luciferase vector by Lipofectamine 2000. Twenty-four hours after transfection, cells were treated with medium alone, 5
    Figure Legend Snippet: Suppression of the Tlr13 gene promoter by LPS and PGN. A , Raw264.7 cells were transfected with Tlr13 promoter p-341 plus Renilla -TK luciferase vector by Lipofectamine 2000. Twenty-four hours after transfection, cells were treated with medium alone, 5

    Techniques Used: Transfection, Luciferase, Plasmid Preparation

    30) Product Images from "Toll-like receptor-mediated activation of neutrophils by influenza A virus"

    Article Title: Toll-like receptor-mediated activation of neutrophils by influenza A virus

    Journal:

    doi: 10.1182/blood-2008-01-132860

    Influenza viral RNA is a ligand for both human TLR7 and TLR8. (A) HEK/TLR8 cells (5 × 10 4 cells) were stimulated with R-848 (3 μM), influenza virus RNA (X31, 0.2 nM) with or without lipofectamine transfection, or IL-1β (100 ng/mL).
    Figure Legend Snippet: Influenza viral RNA is a ligand for both human TLR7 and TLR8. (A) HEK/TLR8 cells (5 × 10 4 cells) were stimulated with R-848 (3 μM), influenza virus RNA (X31, 0.2 nM) with or without lipofectamine transfection, or IL-1β (100 ng/mL).

    Techniques Used: Transfection

    31) Product Images from "Chemotherapy-induced intestinal inflammatory responses are mediated by exosome secretion of double-strand DNA via AIM2 inflammasome activation"

    Article Title: Chemotherapy-induced intestinal inflammatory responses are mediated by exosome secretion of double-strand DNA via AIM2 inflammasome activation

    Journal: Cell Research

    doi: 10.1038/cr.2017.54

    Released dsDNA can be transferred into macrophages via exosomes.  (A)  dsDNA entry into BMMs. HCT-116 cells pre-treated with SN38 (500 nM, 72 h) were stained with DRAQ5 (10 μM) and then co-cultured with BMMs for 24 h more. DRAQ5 staining in BMMs was analysed using flow cytometry.  (B-E)  SN-38-stimulated exosome release in HCT-116 cells. Cells were treated with indicated agents (SN-38, 500 nM; GW4869, 20 μM) for 72 h before harvesting the culture medium.  (B)  Flow chart of culture medium dissection;  (C)  concentrations of dsDNA in each fraction;  (D)  representative transmission electron microscope image (left panel) and size distribution (right panel) based on transmission electron microscope image analysis of exosome fractions. Scale bar, 25 nm;  (E)  immunoblot analysis of CD63 in exosome fractions. The microparticle fraction was used as a control.  (F)  Immunohistochemistry staining of CD63 in ileum isolated from C57BL/6 mice treated with CPT-11 (90 mg/kg, i.p.) for 4 consecutive days. Scale bar, 100 μm.  (G)  Concentrations of dsDNA in culture medium of HCT-116 cells.  (H ,  I)  Localisation of exosome dsDNA in BMMs. Mice BMMs were treated with isolated exosomes from SN-38 treated HCT-116 cells for 2-3 h, and DRAQ5-labelled poly(dA:dT) was transfected into BMMs with Lipofectamine 2000. dsDNA and subcellular organelles were stained using fluorescence confocal microscopy.  (H)  DRAQ5-labelled dsDNA (red), CD11b (green), and nuclei (DAPI, blue).  (I)  DRAQ5-labelled dsDNA (red), lysosomes (LAMP-1, upper, green), early endosomes (EEA1, lower, green), nuclei (blue), and β-actin (purple). The data are representative of three independent experiments and depict the means ± SEM.
    Figure Legend Snippet: Released dsDNA can be transferred into macrophages via exosomes. (A) dsDNA entry into BMMs. HCT-116 cells pre-treated with SN38 (500 nM, 72 h) were stained with DRAQ5 (10 μM) and then co-cultured with BMMs for 24 h more. DRAQ5 staining in BMMs was analysed using flow cytometry. (B-E) SN-38-stimulated exosome release in HCT-116 cells. Cells were treated with indicated agents (SN-38, 500 nM; GW4869, 20 μM) for 72 h before harvesting the culture medium. (B) Flow chart of culture medium dissection; (C) concentrations of dsDNA in each fraction; (D) representative transmission electron microscope image (left panel) and size distribution (right panel) based on transmission electron microscope image analysis of exosome fractions. Scale bar, 25 nm; (E) immunoblot analysis of CD63 in exosome fractions. The microparticle fraction was used as a control. (F) Immunohistochemistry staining of CD63 in ileum isolated from C57BL/6 mice treated with CPT-11 (90 mg/kg, i.p.) for 4 consecutive days. Scale bar, 100 μm. (G) Concentrations of dsDNA in culture medium of HCT-116 cells. (H , I) Localisation of exosome dsDNA in BMMs. Mice BMMs were treated with isolated exosomes from SN-38 treated HCT-116 cells for 2-3 h, and DRAQ5-labelled poly(dA:dT) was transfected into BMMs with Lipofectamine 2000. dsDNA and subcellular organelles were stained using fluorescence confocal microscopy. (H) DRAQ5-labelled dsDNA (red), CD11b (green), and nuclei (DAPI, blue). (I) DRAQ5-labelled dsDNA (red), lysosomes (LAMP-1, upper, green), early endosomes (EEA1, lower, green), nuclei (blue), and β-actin (purple). The data are representative of three independent experiments and depict the means ± SEM.

    Techniques Used: Staining, Cell Culture, Flow Cytometry, Cytometry, Dissection, Transmission Assay, Microscopy, Immunohistochemistry, Isolation, Mouse Assay, Cycling Probe Technology, Transfection, Fluorescence, Confocal Microscopy

    32) Product Images from "Metastasis-associated gene 1 promotes invasion and migration potential of laryngeal squamous cell carcinoma cells"

    Article Title: Metastasis-associated gene 1 promotes invasion and migration potential of laryngeal squamous cell carcinoma cells

    Journal: Oncology Letters

    doi: 10.3892/ol.2013.1729

    RT-PCR determined the effects of pcDNA3- MTA1 , control-siRNA and  MTA1 -siRNA transfection on  MTA1  expression at the mRNA level. pcDNA3- MTA1  and  MTA1 -siRNA were transfected into the cell line with Lipofectamine 2000. Total RNA was extracted and RT-PCR was performed. pcDNA3- MTA1  transfection increased  MTA1  mRNA levels, while  MTA1 -siRNA decreased  MTA1  mRNA levels.  MTA1 , metastasis-associated gene 1.
    Figure Legend Snippet: RT-PCR determined the effects of pcDNA3- MTA1 , control-siRNA and MTA1 -siRNA transfection on MTA1 expression at the mRNA level. pcDNA3- MTA1 and MTA1 -siRNA were transfected into the cell line with Lipofectamine 2000. Total RNA was extracted and RT-PCR was performed. pcDNA3- MTA1 transfection increased MTA1 mRNA levels, while MTA1 -siRNA decreased MTA1 mRNA levels. MTA1 , metastasis-associated gene 1.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Transfection, Expressing

    Western blot analysis of  MTA1  expression after transfection with pcDNA3- MTA1  and pSilencer3.1- MTA1 -siRNA. pcDNA3- MTA1  and  MTA1 -siRNA were transfected into the cell line with Lipofectamine 2000. Whole protein was extracted and loaded for SDS-PAGE separation. The protein was transferred to a nitrocellulose membrane and pcDNA3- MTA1  increased  MTA1  expression and  MTA1 -siRNA greatly decreased  MTA1  levels.  MTA1 , metastasis-associated gene 1.
    Figure Legend Snippet: Western blot analysis of MTA1 expression after transfection with pcDNA3- MTA1 and pSilencer3.1- MTA1 -siRNA. pcDNA3- MTA1 and MTA1 -siRNA were transfected into the cell line with Lipofectamine 2000. Whole protein was extracted and loaded for SDS-PAGE separation. The protein was transferred to a nitrocellulose membrane and pcDNA3- MTA1 increased MTA1 expression and MTA1 -siRNA greatly decreased MTA1 levels. MTA1 , metastasis-associated gene 1.

    Techniques Used: Western Blot, Expressing, Transfection, SDS Page

    33) Product Images from "Glycogen synthase kinase-3 negatively regulates anti-inflammatory interleukin-10 for lipopolysaccharide-induced iNOS/NO biosynthesis and RANTES production in microglial cells"

    Article Title: Glycogen synthase kinase-3 negatively regulates anti-inflammatory interleukin-10 for lipopolysaccharide-induced iNOS/NO biosynthesis and RANTES production in microglial cells

    Journal: Immunology

    doi: 10.1111/j.1365-2567.2008.02959.x

    The effects of glycogen synthase kinase-3 (GSK-3) on nitric oxide (NO), RANTES and interleukin-10 (IL-10) production in lipopolysaccharide (LPS)-stimulated microglia. BV2 mouse microglial cells (2·5 × 10 5 ) were transfected with pcDNA3.1 vector alone (control), pcDNA3.1-GSK-3β wide-type (GSK-3β WT) or dominant-negative form (GSK-3β DN) for 24 hr. (a) After the cells had been treated with LPS (1 μg/ml) for 24 hr, we used Western blotting to determine GSK-3β expression. β-Actin was the internal control. The ratio of GSK-3β to β-actin is shown. (b) We determined NO production using Griess reagent. (c and d) For the 6-hr treatment, we used enzyme-linked immunsorbent assay to determine RANTES and IL-10 production. Cells treated with lipofectamine 2000 were the negative control. The data are means ± SD obtained from three individual experiments. * P
    Figure Legend Snippet: The effects of glycogen synthase kinase-3 (GSK-3) on nitric oxide (NO), RANTES and interleukin-10 (IL-10) production in lipopolysaccharide (LPS)-stimulated microglia. BV2 mouse microglial cells (2·5 × 10 5 ) were transfected with pcDNA3.1 vector alone (control), pcDNA3.1-GSK-3β wide-type (GSK-3β WT) or dominant-negative form (GSK-3β DN) for 24 hr. (a) After the cells had been treated with LPS (1 μg/ml) for 24 hr, we used Western blotting to determine GSK-3β expression. β-Actin was the internal control. The ratio of GSK-3β to β-actin is shown. (b) We determined NO production using Griess reagent. (c and d) For the 6-hr treatment, we used enzyme-linked immunsorbent assay to determine RANTES and IL-10 production. Cells treated with lipofectamine 2000 were the negative control. The data are means ± SD obtained from three individual experiments. * P

    Techniques Used: Transfection, Plasmid Preparation, Dominant Negative Mutation, Western Blot, Expressing, Negative Control

    34) Product Images from "A novel TLR3 inhibitor encoded by African swine fever virus (ASFV)"

    Article Title: A novel TLR3 inhibitor encoded by African swine fever virus (ASFV)

    Journal: Archives of Virology

    doi: 10.1007/s00705-010-0894-7

    I329L inhibits activation of CCL5 and IFN-β promoter. 5A ) I329L inhibits activation of CCL5. HEK-TLR3 cells were transfected with 300 ng empty plasmid vector (pcDNA3) or 300 ng I329L plasmid vector (pcDNA3-I329L) together with 100 ng of CCL5 reporter plasmid vector (CCL5). Six h before harvesting, cells were stimulated extracellularly with 25 μg/ml poly (I:C). 5B ) I329L inhibits activation of IFN-β. HEK-TLR3 cells were transfected with 300 ng empty vector (pcDNA3) or 300 ng of I329L plasmid vector (pcDNA3-I329L) together with 100 ng of IFN-β plasmid vector (IFN-β). Six h before harvesting, cells were stimulated extracellularly with 25 μg/ml poly (I:C). 5C ) I329L does not inhibit intracellular IFN-β activation independently of TLR3. HEK-293T cells were transfected with 300 ng empty vector (pcDNA3) or 300 ng I329L plasmid vector (pcDNA3-I329L) together with 100 ng of IFN-β plasmid reporter (IFN-β). Six h before harvesting, cells were stimulated intracellularly using Lipofectamine 2000 with 0.5 μg/ml poly (I:C). Luciferase activity was normalized to the β-galactosidase activity obtained with the cotransfected β-gal plasmid internal control. Standard deviations are shown by error bars. Details are in Materials and methods
    Figure Legend Snippet: I329L inhibits activation of CCL5 and IFN-β promoter. 5A ) I329L inhibits activation of CCL5. HEK-TLR3 cells were transfected with 300 ng empty plasmid vector (pcDNA3) or 300 ng I329L plasmid vector (pcDNA3-I329L) together with 100 ng of CCL5 reporter plasmid vector (CCL5). Six h before harvesting, cells were stimulated extracellularly with 25 μg/ml poly (I:C). 5B ) I329L inhibits activation of IFN-β. HEK-TLR3 cells were transfected with 300 ng empty vector (pcDNA3) or 300 ng of I329L plasmid vector (pcDNA3-I329L) together with 100 ng of IFN-β plasmid vector (IFN-β). Six h before harvesting, cells were stimulated extracellularly with 25 μg/ml poly (I:C). 5C ) I329L does not inhibit intracellular IFN-β activation independently of TLR3. HEK-293T cells were transfected with 300 ng empty vector (pcDNA3) or 300 ng I329L plasmid vector (pcDNA3-I329L) together with 100 ng of IFN-β plasmid reporter (IFN-β). Six h before harvesting, cells were stimulated intracellularly using Lipofectamine 2000 with 0.5 μg/ml poly (I:C). Luciferase activity was normalized to the β-galactosidase activity obtained with the cotransfected β-gal plasmid internal control. Standard deviations are shown by error bars. Details are in Materials and methods

    Techniques Used: Activation Assay, Transfection, Plasmid Preparation, Luciferase, Activity Assay

    35) Product Images from "A Method to Culture GABAergic Interneurons Derived from the Medial Ganglionic Eminence"

    Article Title: A Method to Culture GABAergic Interneurons Derived from the Medial Ganglionic Eminence

    Journal: Frontiers in Cellular Neuroscience

    doi: 10.3389/fncel.2017.00423

    Setup of transfections of MGE-derived cells. (A,B) Test of different conditions of MGE–derived cells plated on 200 μg/ml PLL and 20 μg/ml LN, transfected at DIV1, and fixed at either DIV4 or DIV6 for immunofluorescence. After transfection cells were cultured without (A) or with 50 ng/ml BDNF (B) . (C) Quantification of the percentage of cells transfected at DIV1 with 0.1 μg of the indicated plasmid DNA and 0.5 μl of Lipofectamine-200, and fixed at different DIVs for immunofluorescence. Bars are means ±SEM of percentage of DAPI-positive cells expressing GFP. (D) Immunofluorescence of a culture transfected with GFP grown in the presence of BDNF, stained with Abs for GFP (green), for MAP2 (red), and with DAPI (blue). (E) DIV6 cells cotrasfected at DIV1 with 0.05 μg of each plasmid (total of 0.1 μg of plasmid DNA), to express GFP and HA-tagged guanine nucleotide exchange factor PIX.
    Figure Legend Snippet: Setup of transfections of MGE-derived cells. (A,B) Test of different conditions of MGE–derived cells plated on 200 μg/ml PLL and 20 μg/ml LN, transfected at DIV1, and fixed at either DIV4 or DIV6 for immunofluorescence. After transfection cells were cultured without (A) or with 50 ng/ml BDNF (B) . (C) Quantification of the percentage of cells transfected at DIV1 with 0.1 μg of the indicated plasmid DNA and 0.5 μl of Lipofectamine-200, and fixed at different DIVs for immunofluorescence. Bars are means ±SEM of percentage of DAPI-positive cells expressing GFP. (D) Immunofluorescence of a culture transfected with GFP grown in the presence of BDNF, stained with Abs for GFP (green), for MAP2 (red), and with DAPI (blue). (E) DIV6 cells cotrasfected at DIV1 with 0.05 μg of each plasmid (total of 0.1 μg of plasmid DNA), to express GFP and HA-tagged guanine nucleotide exchange factor PIX.

    Techniques Used: Transfection, Derivative Assay, Immunofluorescence, Cell Culture, Plasmid Preparation, Expressing, Staining

    36) Product Images from "The Role of p300 Histone Acetyltransferase in UV-Induced Histone Modifications and MMP-1 Gene Transcription"

    Article Title: The Role of p300 Histone Acetyltransferase in UV-Induced Histone Modifications and MMP-1 Gene Transcription

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0004864

    p300 siRNA suppressed UV-induced MMP-1 expression and inhibited UV-enhanced levels of γ-H2AX, p53 and acetyl-H3, and AA inhibited interaction of p300 with γ-H2AX or acetyl-H3 after UV. HDFs were transfected with scrambled control siRNA and p300 siRNA at 100 nM using Lipofectamine as recommended by the manufacturer. A, HDFs were irradiated with UV and incubated at 37°C 24 h after transfection with scrambled control or p300 siRNA. The MMP-1 mRNA level was analyzed by RT-PCR and the amount of MMP-1 protein was analyzed by western blot. B, The effect of p300 siRNA on the protein levels of γ-H2AX, p53, and acetyl-H3 were analyzed by western blotting. C, HDFs were UV-irradiated. After 6 h, whole cell lysates were immunoprecipitated with anti-p300 antibody. D, HDFs were UV-irradiated and post-treated with AA for 6 h. Whole cell lysates were immunoprecipitated with anti-p300 antibody. Proteins were immunoblotted with anti-p300, anti-acetyl-H3, and anti-γ-H2AX antibodies ( upper panels ). Bar graphs ( lower panels ) show quantitative analysis of scanning densitometric values of γ-H2AX, p53, and acetyl-H3 as ratios to β-actin, which was used as a loading control. Data shown are representative of three independent experiments. C: control, UV: UV-irradiated cells, AA: AA treated cells, UV/AA: UV irradiated cells incubated with AA. *P
    Figure Legend Snippet: p300 siRNA suppressed UV-induced MMP-1 expression and inhibited UV-enhanced levels of γ-H2AX, p53 and acetyl-H3, and AA inhibited interaction of p300 with γ-H2AX or acetyl-H3 after UV. HDFs were transfected with scrambled control siRNA and p300 siRNA at 100 nM using Lipofectamine as recommended by the manufacturer. A, HDFs were irradiated with UV and incubated at 37°C 24 h after transfection with scrambled control or p300 siRNA. The MMP-1 mRNA level was analyzed by RT-PCR and the amount of MMP-1 protein was analyzed by western blot. B, The effect of p300 siRNA on the protein levels of γ-H2AX, p53, and acetyl-H3 were analyzed by western blotting. C, HDFs were UV-irradiated. After 6 h, whole cell lysates were immunoprecipitated with anti-p300 antibody. D, HDFs were UV-irradiated and post-treated with AA for 6 h. Whole cell lysates were immunoprecipitated with anti-p300 antibody. Proteins were immunoblotted with anti-p300, anti-acetyl-H3, and anti-γ-H2AX antibodies ( upper panels ). Bar graphs ( lower panels ) show quantitative analysis of scanning densitometric values of γ-H2AX, p53, and acetyl-H3 as ratios to β-actin, which was used as a loading control. Data shown are representative of three independent experiments. C: control, UV: UV-irradiated cells, AA: AA treated cells, UV/AA: UV irradiated cells incubated with AA. *P

    Techniques Used: Expressing, Transfection, Irradiation, Incubation, Reverse Transcription Polymerase Chain Reaction, Western Blot, Immunoprecipitation

    37) Product Images from "Vesicular Trafficking Permits Evasion of cGAS/STING Surveillance During Initial Human Papillomavirus Infection"

    Article Title: Vesicular Trafficking Permits Evasion of cGAS/STING Surveillance During Initial Human Papillomavirus Infection

    Journal: bioRxiv

    doi: 10.1101/2020.03.29.014118

    Bypassing HPV’s Natural Trafficking Pathway Activates cGAS/STING Addition of cationic lipids during infection with translocation-deficient R302/5A mutant HPV16 restores infectivity and allows for cGAS/STING sensing.  (A)  Electron micrograph of HPV16 complexed with the cationic lipid Lipofectamine 2000.  (B)  HaCaTs were infected with 2e8 vge/well of wildtype or R302/5A virion +/- cationic lipid (lipo) for 4 hr and infection was measured by luciferase assay 24h post-infection. While naturally non-infectious, addition of cationic lipids restored infectivity of the R302/5A mutant to levels nearly comparable to those of wildtype HPV16. Statistics calculated by one-way ANOVA ( P
    Figure Legend Snippet: Bypassing HPV’s Natural Trafficking Pathway Activates cGAS/STING Addition of cationic lipids during infection with translocation-deficient R302/5A mutant HPV16 restores infectivity and allows for cGAS/STING sensing. (A) Electron micrograph of HPV16 complexed with the cationic lipid Lipofectamine 2000. (B) HaCaTs were infected with 2e8 vge/well of wildtype or R302/5A virion +/- cationic lipid (lipo) for 4 hr and infection was measured by luciferase assay 24h post-infection. While naturally non-infectious, addition of cationic lipids restored infectivity of the R302/5A mutant to levels nearly comparable to those of wildtype HPV16. Statistics calculated by one-way ANOVA ( P

    Techniques Used: Infection, Translocation Assay, Mutagenesis, Luciferase

    38) Product Images from "HSF1 functions as a transcription regulator for Dp71 expression"

    Article Title: HSF1 functions as a transcription regulator for Dp71 expression

    Journal: Cell Stress & Chaperones

    doi: 10.1007/s12192-014-0558-8

    Over-expression of HSF1 can increase both the Dp71 mRNA and protein expression in HeLa cells. Relative Dp71 mRNA level was analyzed via QRT-PCR after transfection of HSF1-pCDNA3.1(HSF1, vector pcDNA3.1 (Neo), and lipofectamine (Lipo) in HeLa cells ( a
    Figure Legend Snippet: Over-expression of HSF1 can increase both the Dp71 mRNA and protein expression in HeLa cells. Relative Dp71 mRNA level was analyzed via QRT-PCR after transfection of HSF1-pCDNA3.1(HSF1, vector pcDNA3.1 (Neo), and lipofectamine (Lipo) in HeLa cells ( a

    Techniques Used: Over Expression, Expressing, Quantitative RT-PCR, Transfection, Plasmid Preparation

    HSF1 knockdown suppressed the expression of Dp71 in HeLa cells. Relative Dp71 mRNA level in HeLa cell ( a ) was analyzed via QRT-PCR after transfection of psilencer-HSF1 plasmids ( HSF1 ), psilencer plasmid ( Neo ) and lipofectamine ( Lipo ). The Dp71 mRNA expressions
    Figure Legend Snippet: HSF1 knockdown suppressed the expression of Dp71 in HeLa cells. Relative Dp71 mRNA level in HeLa cell ( a ) was analyzed via QRT-PCR after transfection of psilencer-HSF1 plasmids ( HSF1 ), psilencer plasmid ( Neo ) and lipofectamine ( Lipo ). The Dp71 mRNA expressions

    Techniques Used: Expressing, Quantitative RT-PCR, Transfection, Plasmid Preparation

    39) Product Images from "IRF7 in the Australian Black Flying Fox, Pteropus alecto: Evidence for a Unique Expression Pattern and Functional Conservation"

    Article Title: IRF7 in the Australian Black Flying Fox, Pteropus alecto: Evidence for a Unique Expression Pattern and Functional Conservation

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0103875

    qRT-PCR detection of bat IRF7 (A) mRNA expression in P. alecto tissues. Data were normalized with the housekeeping gene 18s rRNA and represent mean results from three individual apparently-healthy wild-caught bats. Error bars represent standard errors (B) Production time course of bat IRF7 following polyI:C stimulation in the bat lung PaLuT02 cell line. Cells were either treated with polyI:C or mixed with Lipofectamine 2000 (transfection) with polyI:C and collected at the indicated time points for RNA extraction and qRT-PCR analysis. The data were normalized against the 18s rRNA gene.
    Figure Legend Snippet: qRT-PCR detection of bat IRF7 (A) mRNA expression in P. alecto tissues. Data were normalized with the housekeeping gene 18s rRNA and represent mean results from three individual apparently-healthy wild-caught bats. Error bars represent standard errors (B) Production time course of bat IRF7 following polyI:C stimulation in the bat lung PaLuT02 cell line. Cells were either treated with polyI:C or mixed with Lipofectamine 2000 (transfection) with polyI:C and collected at the indicated time points for RNA extraction and qRT-PCR analysis. The data were normalized against the 18s rRNA gene.

    Techniques Used: Quantitative RT-PCR, Expressing, Transfection, RNA Extraction

    40) Product Images from "Overexpression of cytoglobin gene inhibits hypoxic injury to SH-SY5Y neuroblastoma cells"

    Article Title: Overexpression of cytoglobin gene inhibits hypoxic injury to SH-SY5Y neuroblastoma cells

    Journal: Neural Regeneration Research

    doi: 10.3969/j.issn.1673-5374.2013.23.010

    Transfected SH-SY5Y-GFP-cytoglobin cells (fluorescence microscopy, × 100). SH-SY5Y cells were transfected with pAcGFP1-C1-cytoglobin using Lipofectamine 2000. The successfully-transfected cells showed green fluorescence. GFP: Green fluorescent protein.
    Figure Legend Snippet: Transfected SH-SY5Y-GFP-cytoglobin cells (fluorescence microscopy, × 100). SH-SY5Y cells were transfected with pAcGFP1-C1-cytoglobin using Lipofectamine 2000. The successfully-transfected cells showed green fluorescence. GFP: Green fluorescent protein.

    Techniques Used: Transfection, Fluorescence, Microscopy

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    Thermo Fisher cell transfections
    Nuclear FGFR1 mediates NGF induced neurite outgrowth and activation of the th gene promoter. ( A ) PC12 cells were transfected with two plasmids, one expressing recombinant FGFR1 or control pcDNA3.1 and the second expressing EGFP. EGFP diffuses throughout the cell permitting visualization of the entire neuritic network. More than 90% of cells co-expressed transfected plasmids as reported in previous studies [29] , [73] . Twenty-four hours after <t>transfection</t> cultures were switched to 1% horse serum medium with or without (control) 50 ng/ml NGF for an additional 10 days, after which the cells were imaged using fluorescent microscopy. Bar length - 100 µm. ( B ) The longest process in an individual transfected cell was measured using ImageJ [24] .* mark comparison to pCDNA3.1 (-NGF) and # to pCNA3.1 (+NGF). Transfection of constitutive nuclear FGFR1(SP−/NLS) increased neurite outgrowth approximately 3-fold (*p
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    Nuclear FGFR1 mediates NGF induced neurite outgrowth and activation of the th gene promoter. ( A ) PC12 cells were transfected with two plasmids, one expressing recombinant FGFR1 or control pcDNA3.1 and the second expressing EGFP. EGFP diffuses throughout the cell permitting visualization of the entire neuritic network. More than 90% of cells co-expressed transfected plasmids as reported in previous studies [29] , [73] . Twenty-four hours after transfection cultures were switched to 1% horse serum medium with or without (control) 50 ng/ml NGF for an additional 10 days, after which the cells were imaged using fluorescent microscopy. Bar length - 100 µm. ( B ) The longest process in an individual transfected cell was measured using ImageJ [24] .* mark comparison to pCDNA3.1 (-NGF) and # to pCNA3.1 (+NGF). Transfection of constitutive nuclear FGFR1(SP−/NLS) increased neurite outgrowth approximately 3-fold (*p

    Journal: PLoS ONE

    Article Title: NGF-Induced Cell Differentiation and Gene Activation Is Mediated by Integrative Nuclear FGFR1 Signaling (INFS)

    doi: 10.1371/journal.pone.0068931

    Figure Lengend Snippet: Nuclear FGFR1 mediates NGF induced neurite outgrowth and activation of the th gene promoter. ( A ) PC12 cells were transfected with two plasmids, one expressing recombinant FGFR1 or control pcDNA3.1 and the second expressing EGFP. EGFP diffuses throughout the cell permitting visualization of the entire neuritic network. More than 90% of cells co-expressed transfected plasmids as reported in previous studies [29] , [73] . Twenty-four hours after transfection cultures were switched to 1% horse serum medium with or without (control) 50 ng/ml NGF for an additional 10 days, after which the cells were imaged using fluorescent microscopy. Bar length - 100 µm. ( B ) The longest process in an individual transfected cell was measured using ImageJ [24] .* mark comparison to pCDNA3.1 (-NGF) and # to pCNA3.1 (+NGF). Transfection of constitutive nuclear FGFR1(SP−/NLS) increased neurite outgrowth approximately 3-fold (*p

    Article Snippet: Cell transfections were performed using Lipofectamine 2000 (Invitrogen).

    Techniques: Activation Assay, Transfection, Expressing, Recombinant, Microscopy

    NGF stimulates FGFR1 binding to NGF-activated genes and cooperates with FGFR1 in activation of Nur-responsive elements - NuRE and NBRE. ( A ) In vivo validation of FGFR1 and Nur binding to an NBRE-like site within intron 1 of the DCX gene. Chromatin was immunoprecipitated with FGFR1, Nur77/Nurr1Ab or IgG in dissected rat brain regions including, the olfactory bulb (OB) which express the dcx gene at high levels [62] , and in the cortex (CX), cerebellum (CB) and ventral midbrain (VM), which express the dcx gene at lower levels. The cyclophilin A gene, which lacks potential Nur-binding NurRE, NBRE or RARE like elements was used as a control. Samples were combined from two rats and the graphs show ΔΔCt means±SEM of triplicate assays. ( B ) PC 12 cells were incubated with or without NGF for 48 hours. ChIP-qPCR was performed with FGFR1 or Nur77/Nurr1 antibodies or control IgG within diverse NGF-activated genes. Similar as in vivo, no binding to the cyclophilinA gene was observed (not shown). Graphs show means of duplicate or triplicate samples from a representative experiment. Similar changes were observed in three separate experiments. ( C,D ) FGFR1 augments Nur77 NurRE- and NBRE-dependent transcription in PC12 cells. Cells were transfected with NurRE-Luc ( C ) or NBRE-Luc ( D ) and either FGFR1(SP−/NLS) or dominant negative FGFR1(TK-) in the presence or absence of Nur77. The amount of transfected DNA was adjusted to 1 µg per well. One day after transfection, PC12 cells were incubated with NGF (50 ng/ml) or without NGF for an additional 24 hours. Results represent the mean ± SEM from 2 experiments with at least three replicates. One-Way ANOVA, LSD: *p

    Journal: PLoS ONE

    Article Title: NGF-Induced Cell Differentiation and Gene Activation Is Mediated by Integrative Nuclear FGFR1 Signaling (INFS)

    doi: 10.1371/journal.pone.0068931

    Figure Lengend Snippet: NGF stimulates FGFR1 binding to NGF-activated genes and cooperates with FGFR1 in activation of Nur-responsive elements - NuRE and NBRE. ( A ) In vivo validation of FGFR1 and Nur binding to an NBRE-like site within intron 1 of the DCX gene. Chromatin was immunoprecipitated with FGFR1, Nur77/Nurr1Ab or IgG in dissected rat brain regions including, the olfactory bulb (OB) which express the dcx gene at high levels [62] , and in the cortex (CX), cerebellum (CB) and ventral midbrain (VM), which express the dcx gene at lower levels. The cyclophilin A gene, which lacks potential Nur-binding NurRE, NBRE or RARE like elements was used as a control. Samples were combined from two rats and the graphs show ΔΔCt means±SEM of triplicate assays. ( B ) PC 12 cells were incubated with or without NGF for 48 hours. ChIP-qPCR was performed with FGFR1 or Nur77/Nurr1 antibodies or control IgG within diverse NGF-activated genes. Similar as in vivo, no binding to the cyclophilinA gene was observed (not shown). Graphs show means of duplicate or triplicate samples from a representative experiment. Similar changes were observed in three separate experiments. ( C,D ) FGFR1 augments Nur77 NurRE- and NBRE-dependent transcription in PC12 cells. Cells were transfected with NurRE-Luc ( C ) or NBRE-Luc ( D ) and either FGFR1(SP−/NLS) or dominant negative FGFR1(TK-) in the presence or absence of Nur77. The amount of transfected DNA was adjusted to 1 µg per well. One day after transfection, PC12 cells were incubated with NGF (50 ng/ml) or without NGF for an additional 24 hours. Results represent the mean ± SEM from 2 experiments with at least three replicates. One-Way ANOVA, LSD: *p

    Article Snippet: Cell transfections were performed using Lipofectamine 2000 (Invitrogen).

    Techniques: Binding Assay, Activation Assay, In Vivo, Immunoprecipitation, Incubation, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Transfection, Dominant Negative Mutation

    NGF accelerates nuclear trafficking of FGFR1 by reducing FGFR1 nuclear export. ( A ) FGFR1-EGFP was transfected into PC12 cells. Twenty-four hours after transfection, cultures were treated with 50 ng/ml NGF and LMB (100 ng/ml in 0.1% v/v ethanol) or ethanol alone (0.1% v/v ethanol) for an additional 48 h and during subsequent imaging. Examples of FGFR1-EGFP expressing cells before and after photo-bleaching are shown. DIC image indicates the nuclear and cytoplasmic regions. About 1/3 nuclear area of PC12 cell was bleached by high intensity laser and 2–3 regions of interest (ROI) intensity were measured. ( B ) Single-exponential analysis of FGFR1-EGFP FLIP regression in cytoplasm and nucleus showed that the diffusion rate of FGFR1-EGFP is affected by NGF treatment in live cells. Individual curves represent means of at least 23 cells. NGF significantly increases the FGFR1-EGFP exchange between nucleus and cytoplasm (half-time decreases) without affecting the FGFR1-EGFP mobile population. ( C ) Single-exponential analysis of FGFR1-EGFP FLIP regression in the cytoplasm shows that NGF facilitates FGFR1-EGFP trafficking between the cytoplasm and nucleus (half-time decreases from 121.5 sec to 89.7 sec; One-way AVOVA, LSD*p

    Journal: PLoS ONE

    Article Title: NGF-Induced Cell Differentiation and Gene Activation Is Mediated by Integrative Nuclear FGFR1 Signaling (INFS)

    doi: 10.1371/journal.pone.0068931

    Figure Lengend Snippet: NGF accelerates nuclear trafficking of FGFR1 by reducing FGFR1 nuclear export. ( A ) FGFR1-EGFP was transfected into PC12 cells. Twenty-four hours after transfection, cultures were treated with 50 ng/ml NGF and LMB (100 ng/ml in 0.1% v/v ethanol) or ethanol alone (0.1% v/v ethanol) for an additional 48 h and during subsequent imaging. Examples of FGFR1-EGFP expressing cells before and after photo-bleaching are shown. DIC image indicates the nuclear and cytoplasmic regions. About 1/3 nuclear area of PC12 cell was bleached by high intensity laser and 2–3 regions of interest (ROI) intensity were measured. ( B ) Single-exponential analysis of FGFR1-EGFP FLIP regression in cytoplasm and nucleus showed that the diffusion rate of FGFR1-EGFP is affected by NGF treatment in live cells. Individual curves represent means of at least 23 cells. NGF significantly increases the FGFR1-EGFP exchange between nucleus and cytoplasm (half-time decreases) without affecting the FGFR1-EGFP mobile population. ( C ) Single-exponential analysis of FGFR1-EGFP FLIP regression in the cytoplasm shows that NGF facilitates FGFR1-EGFP trafficking between the cytoplasm and nucleus (half-time decreases from 121.5 sec to 89.7 sec; One-way AVOVA, LSD*p

    Article Snippet: Cell transfections were performed using Lipofectamine 2000 (Invitrogen).

    Techniques: Transfection, Imaging, Expressing, Diffusion-based Assay, Size-exclusion Chromatography

    Hsp105 overexpression reduces CFTR synthesis but promotes its folding. A and B, HEK cells were co-transfected with wild-type ( A ) or ΔF508 ( B ) CFTR together with a control or Hsp105 ( 105 ) expression plasmid. Cells were lysed 24 h after transfection.

    Journal: The Journal of Biological Chemistry

    Article Title: Human Heat Shock Protein 105/110 kDa (Hsp105/110) Regulates Biogenesis and Quality Control of Misfolded Cystic Fibrosis Transmembrane Conductance Regulator at Multiple Levels *

    doi: 10.1074/jbc.M111.297580

    Figure Lengend Snippet: Hsp105 overexpression reduces CFTR synthesis but promotes its folding. A and B, HEK cells were co-transfected with wild-type ( A ) or ΔF508 ( B ) CFTR together with a control or Hsp105 ( 105 ) expression plasmid. Cells were lysed 24 h after transfection.

    Article Snippet: Cell transfection was performed using Lipofectamine 2000 (Invitrogen) or Effectine (Qiagen, Valencia, CA).

    Techniques: Over Expression, Transfection, Expressing, Plasmid Preparation

    MAP3K3 promotes cell proliferation and the epithelial-mesenchymal transition process. (A) A reverse transcription-quantitative polymerase chain reaction assay demonstrated the efficiency of the overexpression plasmid of MAP3K3 in A549 and H460 cells. (B) The indicated transfection on A549 and H460 cellular viabilities was determined with a Cell Counting Kit-8 assay. A549 and H460 cells were transfected with the indicated combinations of pcDNA3 and MAP3K3 or pri-miR-505 and MAP3K3 or the control group. MAP3K3 overexpression promoted cell viability. NS vs. pri-miR-505 and MAP3K3; ** P

    Journal: International Journal of Molecular Medicine

    Article Title: miR-505 inhibits cell growth and EMT by targeting MAP3K3 through the AKT-NFκB pathway in NSCLC cells

    doi: 10.3892/ijmm.2018.4041

    Figure Lengend Snippet: MAP3K3 promotes cell proliferation and the epithelial-mesenchymal transition process. (A) A reverse transcription-quantitative polymerase chain reaction assay demonstrated the efficiency of the overexpression plasmid of MAP3K3 in A549 and H460 cells. (B) The indicated transfection on A549 and H460 cellular viabilities was determined with a Cell Counting Kit-8 assay. A549 and H460 cells were transfected with the indicated combinations of pcDNA3 and MAP3K3 or pri-miR-505 and MAP3K3 or the control group. MAP3K3 overexpression promoted cell viability. NS vs. pri-miR-505 and MAP3K3; ** P

    Article Snippet: Cell transfection was performed by Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.), according to the manufacturer's protocols.

    Techniques: Real-time Polymerase Chain Reaction, Over Expression, Plasmid Preparation, Transfection, Cell Counting

    14-3-3 protein controls proper SAC inactivation in a γ-subtype-dependent manner. ( a ) Forty-eight hours after transfection with an indicated siRNA, HeLa cells were stained with anti-H3-pSer28 (red) and 4',6-diamidino-2-phenylindole (blue). Immunoblotting with anti-H3-pSer28 and anti-α-tubulin (loading control) are also shown in insets. ( b ) The effect of 14-3-3γ knockdown on cell-cycle progression after DTB synchronization. ( c ) Live-cell imaging analyses using H2B-GFP-expressing HeLa cells. Box-and-whisker plots show the elapsed time that cells spent in mitosis from nuclear envelope breakdown ( t =0) to anaphase onset ( n ≥60 from two independent experiments). ( d ) The effect of 14-3-3γ silencing on KT-associated BubR1. Scale bar, 10 μm. ( e, f ) The effects of BubR1 or Mad2 silencing on mitotic arrest induced by 14-3-3γ depletion. Mitotic indices are shown as mean±s.e.m. from three independent experiments ( n > 200 each). * P

    Journal: Nature Communications

    Article Title: PI 3-kinase-dependent phosphorylation of Plk1-Ser99 promotes association with 14-3-3? and is required for metaphase-anaphase transition

    doi: 10.1038/ncomms2879

    Figure Lengend Snippet: 14-3-3 protein controls proper SAC inactivation in a γ-subtype-dependent manner. ( a ) Forty-eight hours after transfection with an indicated siRNA, HeLa cells were stained with anti-H3-pSer28 (red) and 4',6-diamidino-2-phenylindole (blue). Immunoblotting with anti-H3-pSer28 and anti-α-tubulin (loading control) are also shown in insets. ( b ) The effect of 14-3-3γ knockdown on cell-cycle progression after DTB synchronization. ( c ) Live-cell imaging analyses using H2B-GFP-expressing HeLa cells. Box-and-whisker plots show the elapsed time that cells spent in mitosis from nuclear envelope breakdown ( t =0) to anaphase onset ( n ≥60 from two independent experiments). ( d ) The effect of 14-3-3γ silencing on KT-associated BubR1. Scale bar, 10 μm. ( e, f ) The effects of BubR1 or Mad2 silencing on mitotic arrest induced by 14-3-3γ depletion. Mitotic indices are shown as mean±s.e.m. from three independent experiments ( n > 200 each). * P

    Article Snippet: Plasmid transfection was performed with Lipofectamine LTX and PLUS Reagents according to the manufacturer’s protocol (Invitrogen, Carlsbad, CA).

    Techniques: Transfection, Staining, Live Cell Imaging, Expressing, Whisker Assay